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1
Hiemstra, P. S. "Novel roles of protease inhibitors in infection and inflammation." Biochemical Society Transactions 30, no. 2 (April 1, 2002): 116–20. http://dx.doi.org/10.1042/bst0300116.
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The local balance between proteinase inhibitors and proteinases determines local proteolytic activity. Various studies have demonstrated the importance of serine proteinase inhibitors in regulating the activity of serine proteinases that are released by leucocytes during inflammation. Recently it has been shown that these inhibitors may also display functions that are distinct from those associated with the inhibition of leucocyte-derived proteinases. In this review the results of selected studies focusing on three inhibitors of neutrophil elastase, i.e. α1-proteinase inhibitor, secretory leucocyte proteinase inhibitor and elafin, are presented, with the aim of illustrating their possible involvement in the regulation of inflammation, host defence against infection, tissue repair and extracellular matrix synthesis.
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Sinden, Nicola J., Michael J. Baker, David J. Smith, Jan-Ulrich Kreft, Timothy R. Dafforn, and Robert A. Stockley. "α-1-Antitrypsin variants and the proteinase/antiproteinase imbalance in chronic obstructive pulmonary disease." American Journal of Physiology-Lung Cellular and Molecular Physiology 308, no. 2 (January 15, 2015): L179—L190. http://dx.doi.org/10.1152/ajplung.00179.2014.
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The excessive activities of the serine proteinases neutrophil elastase and proteinase 3 are associated with tissue damage in chronic obstructive pulmonary disease. Reduced concentrations and/or inhibitory efficiency of the main circulating serine proteinase inhibitor α-1-antitrypsin result from point mutations in its gene. In addition, α-2-macroglobulin competes with α-1-antitrypsin for proteinases, and the α-2-macroglobulin-sequestered enzyme can retain its catalytic activity. We have studied how serine proteinases partition between these inhibitors and the effects of α-1-antitrypsin mutations on this partitioning. Subsequently, we have developed a three-dimensional reaction-diffusion model to describe events occurring in the lung interstitium when serine proteinases diffuse from the neutrophil azurophil granule following degranulation and subsequently bind to either α-1-antitrypsin or α-2-macroglobulin. We found that the proteinases remained uninhibited on the order of 0.1 s after release and diffused on the order of 10 μm into the tissue before becoming sequestered. We have shown that proteinases sequestered to α-2-macroglobulin retain their proteolytic activity and that neutrophil elastase complexes with α-2-macroglobulin are able to degrade elastin. Although neutrophil elastase is implicated in the pathophysiology of emphysema, our results highlight a potentially important role for proteinase 3 because of its greater concentration in azurophil granules, its reduced association rate constant with all α-1-antitrypsin variants studied here, its greater diffusion distance, time spent uninhibited following degranulation, and its greater propensity to partition to α-2-macroglobulin where it retains proteolytic activity.
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Otlewski, J., D. Krowarsch, and W. Apostoluk. "Protein inhibitors of serine proteinases." Acta Biochimica Polonica 46, no. 3 (September 30, 1999): 531–65. http://dx.doi.org/10.18388/abp.1999_4128.
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Serine proteinases and their natural protein inhibitors belong to the most intensively studied models of protein-protein recognition. Protein inhibitors do not form a single group but can be divided into about 20 different families. Global structures of proteins representing different inhibitor families are completely different and comprise alpha-helical proteins, beta-sheet proteins, alpha/beta-proteins and different folds of small disulfide-rich proteins. Three different types of inhibitors can be distinguished: canonical (standard mechanism) inhibitors, non-canonical inhibitors, and serpins. The canonical inhibitor binds to the enzyme through the exposed and convex binding loop, which is complementary to the active site of the enzyme. The mechanism of inhibition in this group is consistently very similar and resembles that of an ideal substrate. Non-canonical inhibitors, originating from blood sucking organisms, specifically block enzymes of the blood clotting cascade. The interaction is mediated through inhibitor N-terminus which binds to the proteinase forming a parallel beta-sheet. There are also extensive secondary interactions which provide an additional buried area and contribute significantly to the strength and specificity of recognition. Serpins are major proteinase inhibitors occurring in plasma. Similarly to canonical inhibitors, serpins interact with their target proteinases in a substrate-like manner. However, in the case of serpins, cleavage of a single peptide bond in a flexible and exposed binding loop leads to dramatic structural changes.
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Stepanov, V. M., G. N. Rudenskaya, L. P. Revina, Y. B. Gryaznova, E. N. Lysogorskaya, Filippova IYu, and I. I. Ivanova. "A serine proteinase of an archaebacterium, Halobacterium mediterranei. A homologue of eubacterial subtilisins." Biochemical Journal 285, no. 1 (July 1, 1992): 281–86. http://dx.doi.org/10.1042/bj2850281.
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A homogeneous serine proteinase secreted by the extreme halophilic bacterium Halobacterium mediterranei 1538 was isolated by affinity chromatography on bacitracin-Sepharose with a yield of 48% (260-fold purification). The enzyme reveals an optimum for pyroglutamyl-Ala-Ala-Leu p-nitroanilide hydrolysis at pH 8.0-8.5 (Km 0.14 mM; k(cat). 36.9 s-1). Its activity increases linearly with NaCl concentration over the range 2-5 M. The substrate specificity of the enzyme is comparable with that of secretory subtilisins, the extent of protein degradation approaching that attained with proteinase K. The enzyme has a molecular mass of 41 kDa and a pI of 7.5. The N-terminal sequence of H. mediterranei serine proteinase reveals a 50% identity with that of Thermoactinomyces vulgaris serine proteinases, indicating that the enzyme belongs to the subtilisin family. Hence the serine proteinase secreted by the halophilic bacterium should be considered as a functional analogue, and a structural homologue, of eubacterial serine proteinases (subtilisins).
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Rymerson, Robert T., and Robert P. Bodnaryk. "GUT PROTEINASE ACTIVITY IN INSECT PESTS OF CANOLA." Canadian Entomologist 127, no. 1 (February 1995): 41–48. http://dx.doi.org/10.4039/ent12741-1.
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AbstractThe digestive proteinases of three important pests of canola, Brassica napus L. and B. rapa L., in western Canada were characterized by assessing the proteolytic activity of homogenates of their midguts against azocasein or azoalbumin at various pH levels and in the presence of diagnostic proteinase inhibitors. The midgut of larvae of the bertha armyworm, Mamestra configurata Wlk., had maximum proteolytic activity at pH 10.5 which was inhibited 45–60% by serine proteinase inhibitors such as the soybean trypsin inhibitor. The midgut of larvae of the diamondback moth, Plutella xylostella L., had maximum proteolytic activity at pH 10 which was inhibited 56–75% by serine proteinase inhibitors. The two lepidopterans thus use a serine-like proteinase in digestion. The midgut of adults of the flea beetle, Phyllotreta cruciferae Goeze, exhibited maximum proteolytic activity at pH 5 which was inhibited 33–61% by specific cysteine proteinase inhibitors such as cystatin and trans-epoxysuccinyl-L-leucylamido (4-guanidino)-butane (E-64) and was activated strongly by L-cysteine. Aspartic proteinase inhibitors such as pepstatin A also decreased proteolytic activity by 21–50%. Serine proteinase inhibitors were without effect. Therefore, P. cruciferae appears to use both cysteine- and aspartic-like proteinases in digestion. Cotyledons and first true leaves of canola, B. napus cv. Westar, contained inhibitory activity against serine, cysteine, and aspartic proteinases when tested against bovine trypsin, papain, or porcine pepsin, but the level of antiproteinase activity is insufficient to provide significant resistance against any of these pests.
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Coppedge, B. R., J. M. Jones, G. W. Felton, and F. M. Stephen. "Examination of Midgut Proteinases of the Adult Southern Pine Beetle (Coleoptera: Scolytidae)." Journal of Entomological Science 29, no. 4 (October 1, 1994): 457–65. http://dx.doi.org/10.18474/0749-8004-29.4.457.
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The midgut of adult southern pine beetles, Dendroctonus frontalis Zimmermann (Coleoptera: Scolytidae), contains digestive enzymes with optimal proteolytic activity in vitro near pH 7. General proteinase activity was significantly inhibited by serine and cysteine proteinase class inhibitors, while limited activation by cysteine proteinase class activators was apparent. These results indicate that both cysteine and serine proteinases are present in the adult midgut. The presence of both proteinase classes in adult southern pine beetles coincides with previous studies showing widespread occurrence of these two classes of proteinases as digestive enzymes in midguts of other coleopteran species, but represents one of few beetle species known to possess both proteinase classes simultaneously.
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Rosenthal, P. J., K. Kim, J. H. McKerrow, and J. H. Leech. "Identification of three stage-specific proteinases of Plasmodium falciparum." Journal of Experimental Medicine 166, no. 3 (September 1, 1987): 816–21. http://dx.doi.org/10.1084/jem.166.3.816.
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We have identified and characterized three stage-specific proteinases of Plasmodium falciparum that are active at neutral pH. We analyzed ring-, trophozoite-, schizont-, and merozoite-stage parasites by gelatin substrate PAGE and characterized the identified proteinases with class-specific proteinase inhibitors. No proteinase activity was detected with rings. Trophozoites had a 28 kD proteinase that was inhibited by inhibitors of cysteine proteinases. Mature schizonts had a 35-40 kD proteinase that also was inhibited by cysteine proteinase inhibitors. Merozoite fractions had a 75 kD proteinase that was inhibited by serine proteinase inhibitors. The stage-specific activity of these proteinases and the correlation between the effects of proteinase inhibitors on the isolated enzymes with the effects of the inhibitors on whole parasites suggest potential critical functions for these proteinases in the life cycle of malaria parasites.
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Todorova, V. K., D. P. Knox, and M. W. Kennedy. "Proteinases in the excretory/secretory products (ES) of adult Trichinella spiralis." Parasitology 111, no. 2 (August 1995): 201–8. http://dx.doi.org/10.1017/s0031182000064957.
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SUMMARYAdult Trichinella spiralis were maintained in vitro using defined media and the material excreted/secreted (ES) during this time examined for proteolytic enzyme (proteinase) activity using an azocasein assay and gelatin-substrate gels. Several discrete proteinases in the size range 14–100 kDa were observed with optimal activity at pH 7·5. The use of a class-differentiating panel of proteinase inhibitors indicated that serine proteinases were predominant although some inhibition was evident in the presence of cysteine and metalloproteinase inhibitors. Of a panel of potential natural protein substrates tested, ES proteinases only degraded fibrinogen and plasminogen and degradation was, in part, susceptible to the action of serine, cysteine and aspartyl proteinase inhibitors. In addition, antibody harvested from immune but not normal mice inhibited ES proteinase activity, an observation of relevance to the immunobiology of Trichinosis.
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Ikeda, T. "Involvement of cysteine proteinases in excystment of Paragonimus ohirai metacercariae induced by sodium cholate and A23187." Journal of Helminthology 77, no. 1 (March 2003): 21–26. http://dx.doi.org/10.1079/joh2002144.
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AbstractThe involvement of intrinsic proteinases in the excystment of Paragonimus ohirai metacercariae was studied in in vitro excystment induced by sodium (Na) cholate, a bile salt and A23187, a Ca2+ ionophore. The effects of various proteinase inhibitors on the in vitro excystment were examined and similar inhibitory profiles were obtained. Benzyloxycarbonyl-L-leucyl-L-leucinal (Z-Leu-Leu-H), a cysteine proteinase inhibitor and 4-(2-aminoethyl)-benzenesulfonyl fluoride (Pefabloc SC), a serine proteinase inhibitor completely inhibited excystment, while L-3-carboxy-2,3-trans-epoxypropionyl-leucylamido (4-guanidino)-butane (E-64), a cysteine proteinase inhibitor and leupeptin, a cysteine/serine proteinase inhibitor permitted partial excystment at a lower rate, but inhibited it from proceeding from the partial excystment stage. In secretions released from metacercariae during excystment, proteinase activities detected towards various fluorogenic peptidyl substrates were almost completely inhibited by Z-Leu-Leu-H and E-64, but not by Pefabloc SC. Sodium cholate induced a higher secretion of cysteine proteinases and a higher rate of excystment than A23187. Profiles of cysteine proteinase activities towards five peptidyl substrates detected were markedly different among the two secretions and the lysate of newly excysted juveniles. Newly excysted juveniles released cysteine proteinases with similar activity profiles and levels to metacercariae induced by Na cholate-incubation, whereas the release of cysteine proteinases was reduced compared with metacercariae induced by A23187-incubation. These results provide valuable information about the involvement of intrinsic proteinases in metacercarial excystment.
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Otlewski, J., and D. Krowarsch. "Squash inhibitor family of serine proteinases." Acta Biochimica Polonica 43, no. 3 (September 30, 1996): 431–44. http://dx.doi.org/10.18388/abp.1996_4475.
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Squash inhibitors of serine proteinases form an uniform family of small proteins. They are built of 27-33 amino-acid residues and cross-linked with three disulfide bridges. The reactive site peptide bond (P1-P1') is between residue 5 (Lys, Arg or Leu) and 6 (always Ile). High resolution X-ray structures are available for two squash inhibitors complexed with trypsin. NMR solution structures have also been determined for free inhibitors. The major structural motif is a distorted, triple-stranded antiparallel beta-sheet. A similar folding motif has been recently found in a number of proteins, including: conotoxins from fish-hunting snails, carboxypeptidase inhibitor from potato, kalata B1 polypeptide, and in some growth factors (e.g. nerve growth factor, transforming growth factor beta 2, platelet-derived growth factor). Squash inhibitors are highly stable and rigid proteins. They inhibit a number of serine proteinases: trypsin, plasmin, kallikrein, blood clotting factors: Xa and XIIa, cathepsin G. The inhibition spectrum can be much broadened if specific amino-acid substitutions are introduced, especially at residues which contact proteinase. Squash inhibitors inhibit proteinases via the standard mechanism. According to the mechanism, inhibitors are substrates which exibit at neutral pH a high kcat/K(m) index for hydrolysis and resynthesis of the reactive site, and a low value of the hydrolysis constant.
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Kang, Yoon-Suk, Young-Bae Chun, and Moon-Jae Cho. "Purification and Characterization of proteinase from Ruditapes philippinarum infected with Perkinsus sp." Journal of Medicine and Life Science 1, no. 1 (December 1, 2003): 53–56. http://dx.doi.org/10.22730/jmls.2003.1.1.53.
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In this study, we characterized a proteinase from marine bivalve Ruditapes philippinarum (Manila clam) infected with the protozoan parasite Perkinsus sp. When infected with Perkinsus sp., Manila clam produced a proteinase, which was not appeared in non-infected manila clam and Perkinsus hypnospore. A clear baiid of 45 kDa was detected on 0.2% gelatin containing gel SDS-PAGE. The proteinase was inhibited by phenylmethyl sulfonyl fluoride (PMSF) known as a serine proteinase inhibitor. Using Mucin-Sepharose CL-6B affinity chromatography, proteinases were eluted by Tris-HCl buffer (pH 7.4) containing 20 mM EDTA. Like lectins, N-acetyl-D-galactosamine also could elute proteinases. Based on these results, isolated Ruditapes philippinarum proteinase belonged to the class of serine protease and may have lectin activities or may be associated with lectin
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Owen, C. A., M. A. Campbell, P. L. Sannes, S. S. Boukedes, and E. J. Campbell. "Cell surface-bound elastase and cathepsin G on human neutrophils: a novel, non-oxidative mechanism by which neutrophils focus and preserve catalytic activity of serine proteinases." Journal of Cell Biology 131, no. 3 (November 1, 1995): 775–89. http://dx.doi.org/10.1083/jcb.131.3.775.
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Serine proteinases of human polymorphonuclear neutrophils play an important role in neutrophil-mediated proteolytic events; however, the non-oxidative mechanisms by which the cells can degrade extracellular matrix in the presence of proteinase inhibitors have not been elucidated. Herein, we provide the first report that human neutrophils express persistently active cell surface-bound human leukocyte elastase and cathepsin G on their cell surface. Unstimulated neutrophils have minimal cell surface expression of these enzymes; however, phorbol ester induces a 30-fold increase. While exposure of neutrophils to chemoattractants (fMLP and C5a) stimulates modest (two- to threefold) increases in cell surface expression of serine proteinases, priming with concentrations of lipopolysaccharide as low as 100 fg/ml leads to striking (up to 10-fold) increase in chemoattractant-induced cell surface expression, even in the presence of serum proteins. LPS-primed and fMLP-stimulated neutrophils have approximately 100 ng of cell surface human leukocyte elastase activity per 10(6) cells. Cell surface-bound human leukocyte elastase is catalytically active, yet is remarkably resistant to inhibition by naturally occurring proteinase inhibitors. These data indicate that binding of serine proteinases to the cell surface focuses and preserves their catalytic activity, even in the presence of proteinase inhibitors. Upregulated expression of persistently active cell surface-bound serine proteinases on activated neutrophils provides a novel mechanism to facilitate their egress from the vasculature, penetration of tissue barriers, and recruitment into sites of inflammation. Dysregulation of the cell surface expression of these enzymes has the potential to cause tissue destruction during inflammation.
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Rauber, P., B. Walker, S. Stone, and E. Shaw. "Synthesis of lysine-containing sulphonium salts and their properties as proteinase inhibitors." Biochemical Journal 250, no. 3 (March 15, 1988): 871–76. http://dx.doi.org/10.1042/bj2500871.
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Some sulphonium salts derived from lysine were synthesized with the general structure R-Lys-CH2S+-(alkyl)2. They were examined as inhibitors of the cysteine proteinase clostripain, which has a preference for cleaving peptide bonds at the carboxy group of basic amino acids, and of a number of trypsin-related serine proteinases. Clostripain was irreversibly inactivated by all reagents examined, but in the case of the serine proteinases, depending on the reagent structure, irreversible and reversible inhibitions were observed. These were kinetically characterized.
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Bózner, P., and P. Demeš. "Proteinases inTrichomonas vaginalisandTritrichomonas mobilensisare not exclusively of cysteine type." Parasitology 102, no. 1 (February 1991): 113–15. http://dx.doi.org/10.1017/s0031182000060418.
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SUMMARYHigh molecular weight proteinases ofTrichomonas vaginalis(with apparentMrvalues 142 and > 220 kDa) andTritrichomonas mobilensis(Mr67, 86, 104 and 120 kDa), optimally active at pH 8, were analysed in gelatin-containing polyacrylamide gels. All of these proteinases were resistant to serine-, aspartic- as well as cysteine proteinase inhibitors. Both proteolytic bands inT. vaginalisand two proteinases inT. mobilensis(67 and 104 kDa) were inhibited by EDTA and EGTA suggesting that they belong to the metallo-proteinase class. The 67 kDa proteinase ofT. mobilensiswas inhibited also byo-phenanthroline. The other two bands ofT. mobilensis(86, 120 kDa) were not classified to any proteinase group since they appeared to be resistant to the chelating agents tested in this study.
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Broadway, Roxanne M. "Dietary regulation of serine proteinases that are resistant to serine proteinase inhibitors." Journal of Insect Physiology 43, no. 9 (September 1997): 855–74. http://dx.doi.org/10.1016/s0022-1910(97)00028-0.
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Sever, Natasa, Metka Filipic, Joze Brzin, and Tamara T. Lah. "Effect of Cysteine Proteinase Inhibitors on Murine B16 Melanoma Cell Invasion in vitro." Biological Chemistry 383, no. 5 (May 15, 2002): 839–42. http://dx.doi.org/10.1515/bc.2002.088.
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Abstract Various types of proteinases are implicated in the malignant progression of human and animal tumors. Proteinase inhibitors may therefore be useful as therapeutic agents in antiinvasive and antimetastatic treatment. The aims of this study were (1) to estimate the relative importance of proteinases in B16 cell invasion in vitro using synthetic, classspecific proteinase inhibitors and (2) to assess the inhibitory effect of some naturally occurring cysteine proteinase inhibitors. Serine proteinase inhibitor reduced invasiveness by up to 24%, whereas inhibition of aspartic proteinases reduced invasion by 11%. Synthetic inhibitors of cysteine proteinases markedly impaired invasion: cathepsin B inhibitors, particularly Ca 074Me, inhibited invasion from 20 40%, whereas cathepsin L inhibitor Clik 148 reduced invasion by 11%. The potato cysteine proteinase inhibitor PCPI 8.7 inhibited invasion by 21%, whereas another potato inhibitor, PCPI 6.6, and the mushroom cysteine proteinase inhibitor clitocypin had no effects. As the inhibitors that inhibited cathepsin B were in general more efficient at impairing the invasiveness, we conclude that of the two cysteine proteinases, cathepsin B plays a more important role than cathepsin L in murine melanoma cell invasion.
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BJÖRK, Ingemar, Kerstin NORDLING, Elke RAUB-SEGALL, Ulf HELLMAN, and Steven T. OLSON. "Inactivation of papain by antithrombin due to autolytic digestion: a model of serpin inactivation of cysteine proteinases." Biochemical Journal 335, no. 3 (November 1, 1998): 701–9. http://dx.doi.org/10.1042/bj3350701.
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Cross-class inhibition of cysteine proteinases by serpins differs from serpin inhibition of serine proteinases primarily in that no stable serpin–cysteine proteinase complex can be demonstrated. This difference in reaction mechanism was elucidated by studies of the inactivation of the cysteine proteinases, papain and cathepsin L, by the serpin antithrombin. The two proteinases were inactivated with second-order rate constants of (1.6±0.1)×103 and (8.6±0.4)×102 M-1·s-1 respectively. An antithrombin to papain inactivation stoichiometry of ∼ 3 indicated extensive cleavage of the inhibitor concurrent with enzyme inactivation, a behaviour verified by SDS/PAGE. N-terminal sequence analyses showed cleavage predominantly at the P2–P1 bond, but also at the P2´–P3´ bond of antithrombin. The papain band in SDS/PAGE progressively disappeared on reaction of the enzyme with increasing amounts of antithrombin, but no band representing a stable antithrombin–papain complex appeared. SDS/PAGE with 125I-labelled papain showed that the disappearance of papain was caused by cleavage of the enzyme into small fragments. These results suggest a mechanism in which papain attacks a peptide bond in the reactive-bond loop of antithrombin adjacent to that involved in serine proteinase inhibition. The reaction proceeds, similarly to that between serpins and serine proteinases, to form an inactive acyl-intermediate complex, although with the substrate pathway dominating in the papain reaction. In this complex, papain is highly susceptible to proteolysis and is degraded by still active papain, which greatly decreases the lifetime of the complex and results in liberation of fragmented, inactive enzyme. This model may have relevance also for the inactivation of physiologically or pathologically important cysteine proteinases by serpins.
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Chen, Celine H., and Robert A. Stockley. "Targeting neutrophil serine proteinases in alpha-1 antitrypsin deficiency." Rare Disease and Orphan Drugs Journal 2, no. 4 (2022): 15. http://dx.doi.org/10.20517/rdodj.2022.18.
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Alpha-1 antitrypsin (AAT) is the most abundant irreversible serine proteinase inhibitor in the circulation and plays a major role in protecting lung tissue against destruction from neutrophil serine proteinases. Genetic mutation of AAT leads to reduced circulating levels and AAT deficiency (AATD) which is associated with an increased risk of developing emphysema. This observation suggests that the balance between AAT and neutrophil serine proteinase is crucial in maintaining tissue homoeostasis. In AATD, the overexuberant proteinase activity resulting from inadequate AAT control creates a self-perpetuating inflammatory cycle, driving progressive tissue injury. Re-establishing this physiological balance is therefore critical for preserving lung architecture, function, and abrogating disease progression. Several avenues within this pathophysiological pathway are being explored. This chapter addresses the pathophysiological process, current treatments targeting the pathway, and alternative approaches within the pathway that can potentially mitigate proteinase imbalance.
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Light, Albert, Chester T. Duda, Thomas W. Odorzynski, and William G. I. Moore. "Refolding of serine proteinases." Journal of Cellular Biochemistry 31, no. 1 (1986): 19–26. http://dx.doi.org/10.1002/jcb.240310104.
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Takeuchi, K., H. Wood, and R. T. Swank. "Lysosomal elastase and cathepsin G in beige mice. Neutrophils of beige (Chediak-Higashi) mice selectively lack lysosomal elastase and cathepsin G." Journal of Experimental Medicine 163, no. 3 (March 1, 1986): 665–77. http://dx.doi.org/10.1084/jem.163.3.665.
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A profound decrease in activities of the two lysosomal serine proteinases, elastase, and cathepsin G, was found in neutrophils of four independent beige mutants. Elastase and cathepsin G activities were assayed with the specific synthetic substrates MeO-Suc-Ala-Ala-Pro-Val-MCA and Suc-Ala-Ala-Pro-Phe-pNA, respectively. The defect is intrinsic to cells of beige mice, since transplantation of bone marrow from normal to mutant mice restored normal proteinase activity, and normal mice transplanted with beige marrow produced neutrophils with a deficiency of proteinase activity. The loss of elastase and cathepsin G activity was confirmed by separation of [3H]diisopropylfluorophosphate-labeled proteins on denaturing gels, which also revealed that other serine proteinases are at normal levels in beige neutrophil extracts. The deficiency of lysosomal proteinase activity appears specific, in that four other common neutrophil lysosomal enzymes, plus the spectrum of major neutrophil proteins are not affected by the beige mutation. The deficiency of proteinase activity is likely not the primary genetic alteration of the beige mutation, since more than one proteinase is affected, and heterozygous F1 mice have normal rather than intermediate levels of both proteinases. The lowered proteinase activity may contribute to the high susceptibility of beige mice and Chediak-Higashi patients to infection.
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Wilkinson, David J. "Serpins in cartilage and osteoarthritis: what do we know?" Biochemical Society Transactions 49, no. 2 (April 12, 2021): 1013–26. http://dx.doi.org/10.1042/bst20201231.
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Serpins (serine proteinase inhibitors) are an ancient superfamily of structurally similar proteins, the majority of which use an elegant suicide inhibition mechanism to target serine proteinases. Despite likely evolving from a single common ancestor, the 36 human serpins have established roles regulating diverse biological processes, such as blood coagulation, embryonic development and extracellular matrix (ECM) turnover. Genetic mutations in serpin genes underpin a host of monogenic disorders — collectively termed the ‘serpinopathies’ — but serpin dysregulation has also been shown to drive pathological mechanisms in many common diseases. Osteoarthritis is a degenerative joint disorder, characterised by the progressive destruction of articular cartilage. This breakdown of the cartilage is driven by the metalloproteinases, and it has long been established that an imbalance of metalloproteinases to their inhibitors is of critical importance. More recently, a role for serine proteinases in cartilage destruction is emerging; including the activation of latent matrix metalloproteinases and cell-surface receptors, or direct proteolysis of the ECM. Serpins likely regulate these processes, as well as having roles beyond serine proteinase inhibition. Indeed, serpins are routinely observed to be highly modulated in osteoarthritic tissues and fluids by ‘omic analysis, but despite this, they are largely ignored. Confusing nomenclature and an underappreciation for the role of serine proteinases in osteoarthritis (OA) being the likely causes. In this narrative review, serpin structure, biochemistry and nomenclature are introduced, and for the first time, their putative importance in maintaining joint tissues — as well as their dysregulation in OA — are explored.
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Oppert, B., K. Hartzer, and M. Zuercher. "Digestive proteinases in Lasioderma serricorne (Coleoptera: Anobiidae)." Bulletin of Entomological Research 92, no. 4 (August 2002): 331–36. http://dx.doi.org/10.1079/ber2002179.
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AbstractThe cigarette beetle, Lasioderma serricorne (Fabricius), is a common pest of stored foods. A study of digestive proteinases in L. serricorne was performed to identify potential targets for proteinaceous biopesticides, such as proteinase inhibitors. Optimal casein hydrolysis by luminal proteinases of L. serricorne was in pH 8.5–9.0 buffers, although the pH of luminal contents was slightly acidic. Results from substrate and inhibitor analyses indicated that the primary digestive proteinases were serine proteinases. The most effective inhibitors of caseinolytic hydrolysis were from soybean (both Bowman Birk and Kunitz), with some inhibition by chymostatin, N-TOSYL-L-phenylalanine chloromethyl ketone, and leupeptin. Casein zymogram analysis identified at least eight proteolytic activities. Activity blot analyses indicated one major proteinase activity that hydrolysed the trypsin substrate N-α-benzoyl-L-arginine ρ-nitroanilide, and three major proteinase activities that hydrolysed the chymotrypsin substrate N-succinyl ala-ala-pro-phe ρ-nitroanilide. The absence of cysteine, aspartic, and metallo proteinases in L. serricorne digestion was evidenced by the lack of activation by thiol reagents, alkaline pH optima, and the results from class-specific proteinase inhibitors. The data suggest that protein digestion in L. serricorne is primarily dependent on trypsin- and chymotrypsin-like proteinases.
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Bartlett, J. D., and J. P. Simmer. "Proteinases in Developing Dental Enamel." Critical Reviews in Oral Biology & Medicine 10, no. 4 (July 1999): 425–41. http://dx.doi.org/10.1177/10454411990100040101.
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For almost three decades, proteinases have been known to reside within developing dental enamel. However, identification and characterization of these proteinases have been slow and difficult, because they are present in very small quantities and they are difficult to purify directly from the mineralizing enamel. Enamel matrix proteins such as amelogenin, ameloblastin, and enamelin are cleaved by proteinases soon after they are secreted, and their cleavage products accumulate in the deeper, more mature enamel layers, while the full-length proteins are observed only at the surface. These results suggest that proteinases are necessary for "activating" enamel proteins so the parent proteins and their cleavage products may perform different functions. A novel matrix metalloproteinase named enamelysin (MMP-20) was recently cloned from tooth tissues and was later shown to localize primarily within the most recently formed enamel. Furthermore, recombinant porcine enamelysin was demonstrated to cleave recombinant porcine amelogenin at virtually all of the sites that have previously been described in vivo. Therefore, enamelysin is at least one enzyme that may be important during early enamel development. As enamel development progresses to the later stages, a profound decrease in the enamel protein content is observed. Proteinases have traditionally been assumed to degrade the organic matrix prior to its removal from the enamel. Recently, a novel serine proteinase named enamel matrix serine proteinase-1 (EMSP1) was cloned from enamel organ epithelia. EMSP1 localizes primarily to the early maturation stage enamel and may, therefore, be involved in the degradation of proteins prior to their removal from the maturing enamel. Other, as yet unidentified, proteinases and proteinase inhibitors are almost certainly present within the forming enamel and await discovery.
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Zumbrunn, A., S. Stone, and E. Shaw. "The synthesis and properties of peptidylmethylsulphonium salts with two cationic residues as potential inhibitors of prohormone processing." Biochemical Journal 256, no. 3 (December 15, 1988): 989–94. http://dx.doi.org/10.1042/bj2560989.
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Peptidylmethylsulphonium salts incorporating consecutive basic residues at the C-terminus of the peptidyl portion such as -Arg-Arg-, -Arg-Lys-, -Lys-Lys- and -Lys-Arg- were synthesized and examined as proteinase inhibitors. Serine proteinases with a specificity directed towards hydrolysis at cationic residues were found to be unaffected by these derivatives. On the other hand, cysteine proteinases, cathepsin B and, in particular, clostripain were readily inactivated by affinity labelling. The reagents thus are of promise for the study of prohormone processing promoted by cysteine proteinases.
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Milner, Malgorzata, Jadwiga Chroboczek, and Wlodzimierz Zagorski-Ostoja. "Engineered resistance against proteinases." Acta Biochimica Polonica 54, no. 3 (September 6, 2007): 523–36. http://dx.doi.org/10.18388/abp.2007_3226.
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Exogenous proteinase inhibitors are valuable and economically interesting protective biotechnological tools. We examined whether small proteinase inhibitors when fused to a selected target protein can protect the target from proteolytic degradation without simultaneously affecting the function and activity of the target domain. Two proteinase inhibitors were studied: a Kazal-type silk proteinase inhibitor (SPI2) from Galleria mellonella, and the Cucurbita maxima trypsin inhibitor I (CMTI I). Both inhibitors target serine proteinases, are small proteins with a compact structure stabilized by a network of disulfide bridges, and are expressed as free polypeptides in their natural surroundings. Four constructs were prepared: the gene for either of the inhibitors was ligated to the 5' end of the DNA encoding one or the other of two selected target proteins, the coat protein (CP) of Potato potyvirus Y or the Escherichia coli beta-glucuronidase (GUS). CMTI I fused to the target proteins strongly hampered their functions. Moreover, the inhibitory activity of CMTI I was retained only when it was fused to the CP. In contrast, when fused to SPI2, specific features and functions of both target proteins were retained and the inhibitory activity of SPI2 was fully preserved. Measuring proteolysis in the presence or absence of either inhibitor, we demonstrated that proteinase inhibitors can protect target proteins used either free or as a fusion domain. Interestingly, their inhibitory efficiency was superior to that of a commercial inhibitor of serine proteinases, AEBSF.
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Hornebeck, W., C. Lafuma, L. Robert, M. Móczár, and E. Móczár. "Heparin and its Derivatives Modulate Serine Proteinases (SERPS) Serine Proteinase Inhibitors (SERPINS) Balance." Pathology - Research and Practice 190, no. 9-10 (October 1994): 895–902. http://dx.doi.org/10.1016/s0344-0338(11)80993-3.
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Oozono, Shinji, Nobuhiko Yamauchi, Kyohei Nishimura, Kenji Matsumoto, Ryo Watanabe, Kaiyu Kubota, Shinya Aramaki, et al. "Expression of rat uterine serine proteinases homologous to mouse implantation serine proteinase 2." Journal of Experimental Zoology Part B: Molecular and Developmental Evolution 310B, no. 8 (December 15, 2008): 642–49. http://dx.doi.org/10.1002/jez.b.21237.
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Yang, Haiwei, Tao Li, Jifu Wei, Huiyun Zhang, and Shaoheng He. "Induction of Tumor Necrosis Factor (TNF) Release from Subtypes of T Cells by Agonists of Proteinase Activated Receptors." Mediators of Inflammation 2013 (2013): 1–10. http://dx.doi.org/10.1155/2013/165453.
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Serine proteinases have been recognized as playing an important role in inflammation via proteinase activated receptors (PARs). However, little is known about the influence of serine proteinases and PARs on TNF secretion from highly purified T cells. We challenged T cells from human peripheral blood with serine proteinases and agonist peptides of PARs and measured the levels of TNF in culture supernatants by ELISA. The results showed that thrombin and trypsin, but not tryptase, stimulated approximately up to 2.5-fold increase in TNF release from T cells following 16 h incubation. Proteinase inhibitors and PAR-1 antagonist SCH 79797 almost completely abolished thrombin- and trypsin-induced TNF release from T cells. Agonist peptides of PAR-1, but not PAR-2 induced TNF release from T cells. Moreover, trypsin- and thrombin-induced upregulated expression of TNF was observed in CD4+, IL-4+, or CD25+ T cells, but not in IFN+ or IL-17+ T cells. The signaling pathways MAPK/ERK and PI3K/Akt are involved in the thrombin- and trypsin-induced TNF release from T cells. In conclusion, thrombin and trypsin can induce TNF release from IL-4+ and CD25+ T cells through activation of PAR-1 and therefore contribute to regulation of immune response and inflammation of the body.
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DALTON, J. P., K. A. CLOUGH, M. K. JONES, and P. J. BRINDLEY. "The cysteine proteinases of Schistosoma mansoni cercariae." Parasitology 114, no. 2 (February 1997): 105–12. http://dx.doi.org/10.1017/s003118209600830x.
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Based on substrate preferences, cercariae of Schistosoma mansoni were seen to express both cathepsin L and cathepsin B cysteine proteinases, although the former activity was many -fold greater. Two cathepsin L activities identified in cercarial extracts by zymography co-migrated with activities in extracts of 3 h and 24 h schisotosomula and in extracts of adult worms. Since these enzymes have been implicated in haemoglob in digestion by adult worms, they may perform a similar function in schistosomula. Immunolocalization using scanning electron micrographs showed that cathepsin L and cathepsin B proteinases were present in the cercarial post-acetabular glands. In addition, cercarial serine proteinase activities considered to facilitate skin penetration efficiently cleaved the substrates Z-Gly-Pro-Arg-NHMec and Z-Gly-Pro-Lys-NHMec. Cercariae release most of this serine proteinase activity when induced to secrete the contents of their acetabular glands. In contrast, newly transformed 3 h and 24 h schistosomula did not express this activity.
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Folco, E. J. E., L. Busconi, C. B. Martone, and J. J. Sánchez. "Fish skeletal muscle contains a novel serine proteinase with an unusual subunit composition." Biochemical Journal 263, no. 2 (October 15, 1989): 471–75. http://dx.doi.org/10.1042/bj2630471.
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Proteinase I, an enzyme previously shown to be able to degrade contractile and cytoskeletal elements of white-croaker (Micropogon opercularis) myofibrils, was purified to apparent homogeneity by chromatography on DEAE-Sephacel, octyl-Sepharose CL 4B and arginine-Sepharose 4B. Its Mr was determined to be 269,000 by Sephacryl S-300 gel filtration. Under denaturing conditions, the enzyme dissociated into two subunits with Mr 20,000 and 15,500, in a molar ratio of 1.8:1. Proteinase I showed a pH optimum of 8.5. The enzyme was strongly inhibited by several serine proteinase inhibitors, whereas inhibitors of the other types of proteinases did not affect, or only scarcely affected, its activity. Several N-terminal-blocked 4-methyl-7-coumarylamide substrates having either arginine or lysine residues adjacent to the fluorogenic group were efficiently hydrolysed by the enzyme. These results indicate that proteinase I is a trypsin-like serine proteinase. The enzyme appears to be distinct from other proteinases previously described in skeletal muscle, and might be involved in the catabolism of myofibrillar proteins.
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Talbot, James A., Klaus Nielsen, and Lynette B. Corbeil. "Cleavage of proteins of reproductive secretions by extracellular proteinases of Tritrichomonas foetus." Canadian Journal of Microbiology 37, no. 5 (May 1, 1991): 384–90. http://dx.doi.org/10.1139/m91-062.
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Cleavage of host defense proteins from reproductive secretions was investigated as a potential virulence mechanism for Tritrichomonas foetus extracellular proteinases. Three categories of susceptibility to digestion were found among the defense proteins tested. Cleavage of fibrinogen, fibronectin, and albumin occurred rapidly with more than 50% of these digested within 30 min. Lactoferrin, immunoglobulin G1, and immunoglobulin G2 were more than 50% digested after 4 h. Transferrin, immunoglobulin M, and immunoglobulin A were the most resistant to the Tritrichomonas foetus extracellular proteinases, since 50% or more of the parent molecule remained after 24 h. The responsible proteinases were classified as cysteine (thiol) proteinases because cleavage was inhibited by the cysteine proteinase specific inhibitor, trans-epoxysuccinyl-L-leucylamido-(4-guanidino)butane and not by the serine proteinase specific inhibitor, phenylmethylsulfonyl fluoride. In addition, α2-macro-globulin, but not α1-antitrypsin, inhibits the action of the proteinases. The ratio of this naturally occurring inhibitor to the quantity of proteinases released may determine whether the above substrates are cleaved in vivo. Since these substrates are implicated in iron acquisition, cell adherence, and acquired immunity, Tritrichomonas foetus proteinases are likely to play a role in host–parasite interactions. Key words: cysteine proteinase, Tritrichomonas foetus, host defense proteins, reproductive secretions.
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de Haar, Susanne F., Pieter S. Hiemstra, Martijn T. J. M. van Steenbergen, Vincent Everts, and Wouter Beertsen. "Role of Polymorphonuclear Leukocyte-Derived Serine Proteinases in Defense against Actinobacillus actinomycetemcomitans." Infection and Immunity 74, no. 9 (September 2006): 5284–91. http://dx.doi.org/10.1128/iai.02016-05.
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ABSTRACT Periodontitis is a chronic destructive infection of the tooth-supportive tissues, which is caused by pathogenic bacteria such as Actinobacillus actinomycetemcomitans. A severe form of periodontitis is found in Papillon-Lefèvre syndrome (PLS), an inheritable disease caused by loss-of-function mutations in the cathepsin C gene. Recently, we demonstrated that these patients lack the activity of the polymorphonuclear leukocyte (PMN)-derived serine proteinases elastase, cathepsin G, and proteinase 3. In the present study we identified possible pathways along which serine proteinases may be involved in the defense against A. actinomycetemcomitans. Serine proteinases are capable to convert the PMN-derived hCAP-18 into LL-37, an antimicrobial peptide with activity against A. actinomycetemcomitans. We found that the PMNs of PLS patients released lower levels of LL-37. Furthermore, because of their deficiency in serine proteases, the PMNs of PLS patients were incapable of neutralizing the leukotoxin produced by this pathogen, which resulted in increased cell damage. Finally, the capacity of PMNs from PLS patients to kill A. actinomycetemcomitans in an anaerobic environment, such as that found in the periodontal pocket, seemed to be reduced. Our report demonstrates a mechanism that suggests a direct link between an inheritable defect in PMN functioning and difficulty in coping with a periodontitis-associated pathogen.
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Santos-de-Souza, Raquel, Luzia Monteiro de Castro Côrtes, Karen dos Santos Charret, Léa Cysne-Finkelstein, Carlos Alves, and Franklin Souza-Silva. "Serine Proteinases in Leishmania (Viannia) braziliensis Promastigotes Have Distinct Subcellular Distributions and Expression." International Journal of Molecular Sciences 20, no. 6 (March 15, 2019): 1315. http://dx.doi.org/10.3390/ijms20061315.
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Serine proteinases in Leishmania (Viannia) braziliensis promastigotes were assessed in this work. This study included the investigation of the enzymatic activity of subcellular fractions obtained from benzamidine affinity chromatography, reverse transcription polymerase chain reactions, and in silico assays of subcellular localization of subtilisin. Promastigote serine proteinases showed gelatinolytic activity with molecular masses of 43 kDa to 170 kDa in the cytosolic fraction and 67 kDa to 170 kDa in the membranous fraction. Serine proteinase activities were detected using N-benzyloxycarbonyl-l-phenylalanyl-l-arginine 7-amino-4-methylcoumarin (Z-FR-AMC) and N-succinyl-l-alanine-l-phenylalanine-l-lysine 7-amino-4-methylcoumarin (Suc-AFK-AMC) as substrates in the cytosolic fraction (Z-FR-AMC = 392 ± 30 µmol.min−1 mg of protein−1 and Suc-AFK-AMC = 252 ± 20 µmol.min−1 mg of protein−1) and in the membranous fraction (Z-FR-AMC = 53 ± 5 µmol.min−1 mg of protein−1 and Suc-AFK-AMC = 63.6 ± 6.5 µmol.min−1 mg of protein−1). Enzyme specificity was shown by inhibition with aprotinin (19% to 80% inhibition) and phenylmethanesulfonyl fluoride (3% to 69%), depending on the subcellular fraction and substrate. The expression of subtilisin (LbrM.13.0860 and LbrM.28.2570) and tryparedoxin peroxidase (LbrM.15.1080) genes was observed by the detection of RNA transcripts 200 bp, 162 bp, and 166 bp long, respectively. Subsequent in silico assays showed LbrM.13.0860 can be located in the cytosol and LbrM.28.2570 in the membrane of the parasite. Data obtained here show the subcellular distribution and expression of serine proteinases, including the subtilisin-like serine proteinases in L. (V.) braziliensis promastigotes.
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Stepanov, Valentin M., Galina N. Rudenskaya, Lyudmila I. Vasil'eva, Irina N. Krest'anova, Olga M. Khodova, and Yurii E. Bartoshevitch. "Serine proteinases from Acremonium chrysogenum." International Journal of Biochemistry 18, no. 4 (January 1986): 369–75. http://dx.doi.org/10.1016/0020-711x(86)90043-1.
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Malcolm, Bruce A. "The picornaviral 3C proteinases: Cysteine nucleophiles in serine proteinase folds." Protein Science 4, no. 8 (August 1995): 1439–45. http://dx.doi.org/10.1002/pro.5560040801.
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González-Páez, Gonzalo Ernesto, Fernando Alba-Hurtado, Carlos Gerardo García-Tovar, and Raúl Argüello-García. "Proteinases in Excretory-Secretory Products ofToxocara canisSecond-Stage Larvae: Zymography and Modeling Insights." BioMed Research International 2014 (2014): 1–9. http://dx.doi.org/10.1155/2014/418708.
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Components released in excretory-secretory products ofToxocara canislarvae (TES) include phosphatidylethanolamine-binding proteins (TES26), mucins (TES120, MUC2-5), and C-type lectins (TES32, TES70) and their biochemical, immunological, and diagnostic properties have been extensively studied albeit proteinase activities towards physiological substrates are almost unknown. Proteolytic activities in TES samples were first analyzed by gel electrophoresis with gelatin as substrate. Major activities of ~400, 120, and 32 kDa in TES were relatively similar over a broad pH range (5.5–9.0) and all these were of the serine-type as leupeptin abolished gelatinolysis. Further, the ~400 kDa component degraded all physiological substrates tested (laminin, fibronectin, albumin, and goat IgG) and the 120 kDa component degraded albumin and goat IgG while proteinases of lower MW (45, 32, and 26 kDa) only degraded laminin and fibronectin, preferentially at alkaline pH (9.0). By protein modeling approaches using the known sequences of TES components, only TES26 and MUC4 displayed folding patterns significantly related to reference serine proteinases. These data suggest that most of serine proteinase activities secretedin vitroby infective larvae ofT. canishave intriguing nature but otherwise help the parasite to affect multiple components of somatic organs and bodily fluids within the infected host.
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Lüdemann, J., B. Utecht, and W. L. Gross. "Anti-neutrophil cytoplasm antibodies in Wegener's granulomatosis recognize an elastinolytic enzyme." Journal of Experimental Medicine 171, no. 1 (January 1, 1990): 357–62. http://dx.doi.org/10.1084/jem.171.1.357.
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The target antigen of anti-neutrophil cytoplasm antibodies (ACPA; also known as ANCA) was isolated by affinity chromatography from supernatants of human neutrophils, stimulated with phorbol ester to induce degranulation. Sequence analysis of the antigen revealed 17 NH2-terminal amino acids (IVGGHEAQPHIRPIYMA), which have considerable sequence homology with known serine proteinases. Investigation of the enzymatic activity showed that the antigen is a neutral serine proteinase that is able to cleave elastin. Since the molecular weight of the antigen, its substrate specificity, and its inhibitor profile reported in this study are identical with those reported recently for proteinase 3, we conclude that ACPA are most probably directed against proteinase 3.
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Schechter, N. M., A. M. Irani, J. L. Sprows, J. Abernethy, B. Wintroub, and L. B. Schwartz. "Identification of a cathepsin G-like proteinase in the MCTC type of human mast cell." Journal of Immunology 145, no. 8 (October 15, 1990): 2652–61. http://dx.doi.org/10.4049/jimmunol.145.8.2652.
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Abstract Human mast cells can be divided into two subsets based on serine proteinase composition: a subset that contains the serine proteinases tryptase and chymase (MCTC), and a subset that contains only tryptase (MCT). In this study we examined both types of mast cells for two additional proteinases, cathepsin G and elastase, which are the major serine proteinases of neutrophils. Because human mast cell chymase and cathepsin G are both chymotrypsin-like proteinases, the properties of these enzymes were further defined to confirm their distinctiveness. Comparison of their N-terminal sequences showed 30% nonidentity over the first 35 amino acids, and comparison of their amino acid compositions demonstrated a marked difference in their Arg/Lys ratios, which was approximately 1 for chymase and 10 for cathepsin G. Endoglycosidase F treatment increased the electrophoretic mobility of chymase on SDS gels, indicating significant N-linked carbohydrate on chymase; no effect was observed on cathepsin G. Immunoprecipitation and immunoblotting with specific antisera to each proteinase revealed little, if any, detectable cross-reactivity. Immunocytochemical studies showed selective labelling of MCTC type mast cells by cathepsin G antiserum in sections of human skin, lung, and bowel. No labeling of mast cells by elastase antiserum was detected in the same tissues, or in dispersed mast cells from lung and skin. A protein cross-reactive with cathepsin G was identified in extracts of human skin mast cells by immunoblot analysis. This protein had a slightly higher Mr (30,000) than the predominant form of neutrophil cathepsin G (Mr 28,000), and could not be separated from chymase (Mr 30,000) by SDS gel electrophoresis because of the size similarity. Using casein, a protein substrate hydrolyzed at comparable rates by chymase and cathepsin G, it was shown that about 30% of the caseinolytic activity in mast cell extracts was sensitive to inhibitors of cathepsin G that had no effect on chymase. Hydrolytic activity characteristic of elastase was not detected in these extracts. These studies indicate that human MCTC mast cells may contain two different chymotrypsin-like proteinases: chymase and a proteinase more closely related to cathepsin G, both of which are undetectable in MCT mast cells. Neutrophil elastase, on the other hand, was not detected in human mast cells by our procedures.
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Böhm, B., R. Deutzmann, and H. Burkhardt. "Purification of a serine-proteinase inhibitor from human articular cartilage. Identity with the acid-stable proteinase inhibitor of mucous secretions." Biochemical Journal 274, no. 1 (February 15, 1991): 269–73. http://dx.doi.org/10.1042/bj2740269.
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An inhibitor of the serine proteinases human leucocyte elastase (EC 3.4.21.37), of cathepsin G (EC 3.4.21.20) and of trypsin (EC 3.4.21.4) has been purified from human articular cartilage. The apparent Mr of the cationic (pI greater than 10) protein was determined to 15,000 by SDS/PAGE. It was shown to cross-react in Western blot with a specific antibody to a recombinant-derived serine-proteinase inhibitor of human mucous secretions. Identity of both inhibitors is indicated by the determination of the N-terminal amino acid sequence of the cartilage-derived serine-proteinase inhibitor. In all 24 residues the cartilage inhibitor was shown to be identical with the human secretory leucocyte proteinase inhibitor (‘SLPI’). The inhibitor molecule may play a crucial role in the protection of cartilage matrix proteins against proteolytic attack.
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Vissers, M. C. M., and C. C. Winterbourn. "Activation of human neutrophil gelatinase by endogenous serine proteinases." Biochemical Journal 249, no. 2 (January 15, 1988): 327–31. http://dx.doi.org/10.1042/bj2490327.
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The role of serine proteinases and oxidants in the activation of gelatinase released from human neutrophils was investigated. Gelatinase was measured by its ability to degrade both gelatin and native glomerular basement-membrane type IV collagen. When fMet-Leu-Phe or phorbol 12-myristate 13-acetate was used to stimulate the neutrophils, no gelatinase activity was measured in the absence of a mercurial activator, indicating that the enzyme was released entirely in latent form. However, when fMet-Leu-Phe-stimulated cells were treated with cytochalasin B, 50-70% of the maximal gelatinase activity was released. Activation was blocked by the serine-proteinase inhibitor phenylmethanesulphonyl fluoride and a specific inhibitor of neutrophil elastase, but was not affected by an inhibitor of cathepsin G. Addition of catalase or azide to prevent oxidative reactions did not affect activation of gelatinase under any conditions of stimulation, indicating that oxidants were not involved in activation. Our results imply that oxidative activation of gelatinase does not occur readily. However, neutrophil serine proteinases, particularly elastase, provide an alternative and apparently more efficient mechanism of activation.
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Oppert, B., P. Walters, and M. Zuercher. "Digestive proteinases of the larger black flour beetle, Cynaeus angustus (Coleoptera: Tenebrionidae)." Bulletin of Entomological Research 96, no. 2 (April 2006): 167–72. http://dx.doi.org/10.1079/ber2005413.
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AbstractDigestion in the larger black flour beetle, Cynaeus angustus (LeConte), was studied to identify new control methods for this pest of stored grains and grain products. The physiological pH of the larval gut, as measured with extracts in water, was approximately 6.1, and the pH for optimal hydrolysis of casein by gut extracts was 6.2 when buffers were reducing. However, under non-reducing conditions, hydrolysis of casein and synthetic serine proteinase substrates was optimal in alkaline buffer. Three major proteinase activities were observed in zymograms using casein or gelatin. Caseinolytic activity of C. angustus gut extracts was inhibited by inhibitors that target aspartic and serine proteinase classes, with minor inhibition by a cysteine proteinase inhibitor. In particular, soybean trypsin and trypsin/chymotrypsin inhibitors were most effective in reducing the in vitro caseinolytic activity of gut extracts. Based on these data, further studies are suggested on the effects of dietary soybean inhibitors of serine proteinases, singly and in combination with aspartic and cysteine proteinase inhibitors, on C. angustus larvae. Results from these studies can be used to develop new control strategies to prevent damage to grains and stored products by C. angustus and similar coleopteran pests.
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Black, M. T., J. G. R. Munn, and A. E. Allsop. "On the catalytic mechanism of prokaryotic leader peptidase 1." Biochemical Journal 282, no. 2 (March 1, 1992): 539–43. http://dx.doi.org/10.1042/bj2820539.
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The catalytic mechanism of leader peptidase 1 (LP1) of the bacterium Escherichia coli has been investigated by a combination of site-directed mutagenesis, assays of enzyme activity in vivo utilizing a strain of E. coli which has a conditional defect in LP1 activity, and gene cloning. The biological activity of mutant forms of E. coli LP1 demonstrates that this enzyme belongs to a novel class of proteinases. The possibility that LP1 may be an aspartyl proteinase has been excluded on the basis of primary sequence comparison and mutagenesis. Assignment of LP1 to one of the other three recognized classes of proteinases (metalloproteinases, thiol proteinases and the classical serine proteinases) can also be excluded, as it is clearly demonstrated that none of the histidine or cysteine residues within LP1 are required for catalytic activity. The Pseudomonas fluorescens lep gene has been cloned and sequenced and the corresponding amino acid sequence compared with that of E. coli LP1. The E. coli LP1 and P. fluorescens LP1 primary sequences are 50% identical after insertion of gaps. The P. fluorescens LP1 has 39 fewer amino acids, a calculated molecular mass of 31903 Da and functions effectively in vivo in E. coli. None of the cysteine residues and only one of the histidine residues which are present in E. coli LP1 are conserved in sequence position in the P. fluorescens LP1 enzyme. The possibility that LP1 is a novel type of serine proteinase is discussed.
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Ge, Zhao-Yu, Pin-Jun Wan, and Zhao-Jun Han. "Cloning and characterization of trypsin- and chymotrypsin-like genes in the striped rice stem borer, Chilo suppressalis." Genome 55, no. 4 (April 2012): 281–88. http://dx.doi.org/10.1139/g2012-015.
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Serine proteinases including trypsins and chymotrypsins play various important roles in insects, including food digestion, immune defense, and zymogen activation. Studies on insect serine proteinases could reveal their feeding preference (polyphagous and monophagous) and facilitate identification of protease inhibitors, which can be engineered for pest management. In this paper, 11 transcripts encoding trypsin- and chymotrypsin-like proteins were cloned from the striped rice stem borer, Chilo suppressalis (Walker). All the predicted proteins share high sequence similarity with known trypsin- and chymotrypsin-like proteins from either lepidopterans or dipterans, and most of the proteins have conserved motifs that are characteristics of serine proteinases. Among the 11 cloned genes, six were expressed predominantly and one exclusively in the midgut of the insect, three were expressed relatively evenly in examined tissues, and one was not expressed in either the gut or hemolymph based on RT–PCR results. The seven genes that were predominantly or exclusively expressed in the gut were also affected by feeding on different host plants. The genes that were expressed in the gut and were affected by host plants are likely to encode digestive proteinases. The identification of trypsin- and chymotrypsin-like genes in this insect species is the first step towards further comparative studies and for identification of insect-specific proteinase inhibitors, which might be engineered to protect rice plants against the striped rice stem borer, which is one of the destructive pests of rice.
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Polanowski, Antoni, Anna Wilimowska-Pelc, Jolanta Kowalska, Joanna Grybel, Monika Zelazko, and Tadeusz Wilusz. "Non-conventional affinity chromatography of serine proteinases and their inhibitors." Acta Biochimica Polonica 50, no. 3 (September 30, 2003): 765–73. http://dx.doi.org/10.18388/abp.2003_3667.
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From among a wide variety of protein purification techniques affinity chromatography has proved to be particularly effective for separation of proteolytic enzymes and their inhibitors. In this article, following a general description of affinity adsorbents used for purification of proteinases, we overview a simple separation procedure for some serine proteinases and their inhibitors by way of affinity chromatography in the presence of high NaCl concentration. It has been shown that some highly specific trypsin inhibitors exhibit also antichymotrypsin activity when high concentration of Na(+) but not K(+) or Li(+) ions are present in the reaction mixture. Taking advantage of this phenomenon the virgin forms of trypsin inhibitors from squash seeds, Kazal-type inhibitor from porcine pancreas and alpha(1)-proteinase inhibitor from human and sheep plasma, as an example, were separated using immobilized chymotrypsin or its inactive derivative methylchymotrypsin in the presence of 5 M NaCl.
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Overall, C. M., and H. Limeback. "Identification and characterization of enamel proteinases isolated from developing enamel. Amelogeninolytic serine proteinases are associated with enamel maturation in pig." Biochemical Journal 256, no. 3 (December 15, 1988): 965–72. http://dx.doi.org/10.1042/bj2560965.
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During tooth formation nearly all of the protein matrix of enamel is removed before final mineralization. To study this process, enamel proteins and proteinases were extracted from pig enamel at different stages of tooth development. In the enamel maturation zones, the major enamel matrix proteins, the amelogenins, were rapidly processed and removed. Possibly associated with this process in vivo are two groups of proteinases which were identified in the enamel extracts by enzymography using amelogenin-substrate and gelatin-substrate polyacrylamide gels and by the degradation in vitro of guanidinium chloride-extracted amelogenins. One group of proteinases with gelatinolytic activity consisted of several neutral metalloendoproteinases having Mr values from 62,000 to 130,000. These proteinases were inactive against amelogenins, casein and albumin, and were present in approximately equal proportions in enamel at all developmental stages. In the other group, two serine proteinases, with apparent non-reduced Mr of 31,000 and 36,000 exhibited amelogeninolytic activity. The substrate preference of the enamel serine proteinases was indicated by their limited degradation of casein and their inability to degrade gelatin and albumin. Contrasting with the distribution of the metalloendoproteinase enzymes, the serine proteinases were found only in the enamel scrapings taken from late-maturing enamel. The amelogenin degradation patterns in vivo, observed in the enamel scrapings, were similar to those produced in assays in vitro using partially purified fractions of enamel proteinases and amelogenin substrate. Together, these data strongly indicate an important role for the serine proteinases, and possibly the gelatinolytic proteinases, in the organized processing of the enamel protein matrix during enamel formation.
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Hawthorne, S. J., D. W. Halton, and B. Walker. "Identification and characterization of the cysteine and serine proteinases of the trematode,Haplometra cylindraceaand determination of their haemoglobinase activity." Parasitology 108, no. 5 (June 1994): 595–601. http://dx.doi.org/10.1017/s0031182000077465.
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SummaryThe excreted/secreted proteinases ofHaplometra cylindraceamaintainedin vitro, were found to hydrolyse the fluorogenic substrates, Z-ArgArg-NHMec and Z-PheArg-NHMec. This activity was shown to be typically that of cysteine proteinases, as turn-over of both substrates could be blocked by pre-incubation with peptidyl diazomethyl ketones. The biotinylated affinity reagent, biotin-Phe Ala-DMK, used in combination with Z-PheTyr(OBut)-DMK, was employed for the labelling and characterization of these cysteine proteinase activities. Three cathepsin B-like species were detected, with molecular weights of 48, 22–23 and 14 kDa, together with a cathepsin L-like enzyme, with a molecular weight of 55 kDa. The proteinases were also found to have hydrolytic activity towards the substrate, Z-GlyGlyArg-NHMec, which could be blocked by pre-incubation with either of the serine proteinase-selective reagents, Z-ArgP(OPh)2, or biotin-LysP(OPh)2, showing the activity to be trypsin-like. Using the biotinylated affinity label to characterize the trypsin-like enzymes revealed two molecular species with molecular weights of 20 and 24 kDa.
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Witko-Sarsat, V., and B. Descamps-Latscha. "Neutrophil-derived Oxidants and Proteinases as Immunomodulatory Mediators in Inflammation." Mediators of Inflammation 3, no. 4 (1994): 257–73. http://dx.doi.org/10.1155/s0962935194000360.
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Neutrophils generate potent microbicidal molecules via the oxygen-dependent pathway, leading to the generation of reactive oxygen intermediates (ROI), and via the non-oxygen dependent pathway, consisting in the release of serine proteinases and metalloproteinases stored in granules. Over the past years, the concept has emerged that both ROI and proteinases can be viewed as mediators able to modulate neutrophil responses as well as the whole inflammatory process. This is well illustrated by the oxidative regulation of proteinase activity showing that oxidants and proteinases acts is concert to optimize the microbicidal activity and to damage host tissues. ROI and proteinases can modify the activity of several proteins involved in the control of inflammatory process. Among them, tumour necrosis factor-α and interleukin-8, are elective targets for such a modulation. Moreover, ROI and proteinases are also able to modulate the adhesion process of neutrophils to endothelial cells, which is a critical step in the inflammatory process.
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Serbin, A., Y. Komar, T. Koval, O. Kharchenko, and T. Andriychuk. "Study of proteolytic activity in rats kidney and liver during the development of chronic alcoholic intoxication." Bulletin of Taras Shevchenko National University of Kyiv. Series: Biology 85, no. 2 (2021): 42–46. http://dx.doi.org/10.17721/1728_2748.2021.85.42-46.
Full textAbstract:
The study of biochemical processes in chronic alcohol intoxication is a topical issue of our time. A deeper understanding of the mechanism of action of proteinases in this pathological condition will help in the development and search for non-invasive diagnostic methods, thereby minimizing the risk of harming human health during complex diagnostic procedures. In our experiment, we investigated the general proteolytic activity, the activity of metalloproteinases and serine proteinases in the liver and kidneys of rats on an experimental model of chronic alcohol intoxication on days 1, 3, 7, and 11 of the experiment, as well as on days 21 and 28 after the cessation of ethanol administration. Male rats weighing 180–200 g were modeled for chronic alcohol intoxication by intragastric administration of 30% ethyl alcohol solution for 10 days on an empty stomach, at the rate of 2 ml per 100 g of animal weight. Liver and kidney homogenate by well-known methods. The concentration was determined by the Bradford method. The total proteolytic activity, the activity of metalloproteinases and serine proteinases were determined by the caseinolytic method with modifications. The total proteolytic activity, the activity of metalloproteinases and serine proteinases were determined by the caseinolytic method with modifications. It was shown that on the 3rd, 7th and 11th days of the experiment in the liver there was an increase in the total proteolytic activity and the activity of metalloproteinases. The activity of serine proteinases significantly increased on days 3 and 7 of the study. In the kidneys, a significant increase in all studied activities was observed only on the 3rd day. Such differences in the activities of metalloproteinases and serine proteinases can be associated with the different roles of these enzymes in physiological processes. Thus, we observed an increase in the activity of serine proteinases in acute intoxication, and in metalloproteinases in chronic intoxication.
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Perkins, S. J., and K. F. Smith. "Identity of the putative serine-proteinase fold in proteins of the complement system with nine relevant crystal structures." Biochemical Journal 295, no. 1 (October 1, 1993): 109–14. http://dx.doi.org/10.1042/bj2950109.
Full textAbstract:
The serine-proteinase domain is responsible for the proteolytic events that occur during complement activation. The sequences of nine serine proteinases of known crystal structure were compared with the serine-proteinase sequences in the six complement proteins C1r, C1s, C2, factor B, factor I and factor D to assess the degree of structural homology of the latter with the crystal structures. All sequence insertions and deletions were readily located at the protein surface. The internal location of disulphide bridges and the surface location of putative glycosylation sites are compatible with this structure. Secondary-structure predictions for the sequences were fully consistent with the crystal structures. It is concluded that the double subdomain beta-sheet motif is retained in the complement sequences, but that localized differences are observed for factor I, C2 and factor B.
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Turner, A. Denise, S. V. Pizzo, George Rozakis, and Ned A. Porter. "Photoreactivation of irreversibly inhibited serine proteinases." Journal of the American Chemical Society 110, no. 1 (January 1988): 244–50. http://dx.doi.org/10.1021/ja00209a040.
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