scholarly journals The primary elastase inhibitor (elastasin) and trypsin inhibitor (contrapsin) in the goat are serpins related to human α1-anti-chymotrypsin

1995 ◽  
Vol 306 (1) ◽  
pp. 191-197 ◽  
Author(s):  
J Potempa ◽  
J J Enghild ◽  
J Travis
Keyword(s):  
Trypsin Inhibitor ◽  
Neutrophil Elastase ◽  
Proteinase Inhibitors ◽  
Specific Inhibitor ◽  
Serine Proteinase ◽  
Peptide Bond ◽  
Reactive Site ◽  
Elastase Inhibitor ◽  
Efficient Inhibitor ◽  
Inhibitory Specificity

Two primary serine proteinase inhibitors in goat plasma have been isolated and characterized. The N-terminal sequence analysis of the purified proteins revealed that they are closely related to each other and are highly homologous to human alpha 1-anti-chymotrypsin rather than alpha 1-proteinase inhibitor. However, despite structural similarities the inhibitory specificity of the goat inhibitors differed from each other and from that of anti-chymotrypsin. In contrast with human anti-chymotrypsin, one of the goat inhibitors was shown to be a strong and specific inhibitor of trypsin (k(ass.) = 1.9 x 10(6) M-1.s-1), whereas the other was an efficient inhibitor of neutrophil elastase (k(ass.) = 1.5 x 10(6) M-1.S-1). Differences in the inhibitory specificity of each protein could readily be attributed to the amino acid sequence within the reactive site region. The trypsin inhibitor with an assumed arginine residue at the P1 position of the reactive-site peptide bond is referred to as ‘contrapsin’, and indicates that the occurrence of contrapsins is not restricted to rodents. In contrast, the inhibitory specificity, resistance to oxidative and proteolytic inactivation and the presence of a P1 leucine residue in the elastase inhibitor is unique among inhibitory serpins that have been characterized to date. Because this serpin is apparently the major elastase inhibitor in goat plasma, it is likely to be involved in the control of goat neutrophil elastase. Therefore, we suggest the name ‘elastasin’, and extend it to any other anti-chymotrypsin related serpins possessing neutrophil-elastase- inhibitory activity.

scholarly journals A Retrospective Analysis of the Cartilage Kunitz Protease Inhibitory Proteins Identifies These as Members of the Inter-α-Trypsin Inhibitor Superfamily with Potential Roles in the Protection of the Articulatory Surface

2019 ◽  
Vol 20 (3) ◽  
pp. 497 ◽  
Author(s):  
Susan Smith ◽  
James Melrose
Keyword(s):  
Articular Cartilage ◽  
Affinity Chromatography ◽  
Trypsin Inhibitor ◽  
Concanavalin A ◽  
Inhibitory Activity ◽  
Proteinase Inhibitors ◽  
Serine Proteinase ◽  
Lectin Affinity Chromatography ◽  
Joint Function ◽  
Lectin Affinity

Aim: The aim of this study was to assess if the ovine articular cartilage serine proteinase inhibitors (SPIs) were related to the Kunitz inter-α-trypsin inhibitor (ITI) family. Methods: Ovine articular cartilage was finely diced and extracted in 6 M urea and SPIs isolated by sequential anion exchange, HA affinity and Sephadex G100 gel permeation chromatography. Selected samples were also subjected to chymotrypsin and concanavalin-A affinity chromatography. Eluant fractions from these isolation steps were monitored for protein and trypsin inhibitory activity. Inhibitory fractions were assessed by affinity blotting using biotinylated trypsin to detect SPIs and by Western blotting using antibodies to α1-microglobulin, bikunin, TSG-6 and 2-B-6 (+) CS epitope generated by chondroitinase-ABC digestion. Results: 2-B-6 (+) positive 250, 220,120, 58 and 36 kDa SPIs were detected. The 58 kDa SPI contained α1-microglobulin, bikunin and chondroitin-4-sulfate stub epitope consistent with an identity of α1-microglobulin-bikunin (AMBP) precursor and was also isolated by concanavalin-A lectin affinity chromatography indicating it had N-glycosylation. Kunitz protease inhibitor (KPI) species of 36, 26, 12 and 6 kDa were autolytically generated by prolonged storage of the 120 and 58 kDa SPIs; chymotrypsin affinity chromatography generated the 6 kDa SPI. KPI domain 1 and 2 SPIs were separated by concanavalin lectin affinity chromatography, domain 1 displayed affinity for this lectin indicating it had N-glycosylation. KPI 1 and 2 displayed potent inhibitory activity against trypsin, chymotrypsin, kallikrein, leucocyte elastase and cathepsin G. Localisation of versican, lubricin and hyaluronan (HA) in the surface regions of articular cartilage represented probable binding sites for the ITI serine proteinase inhibitors (SPIs) which may preserve articulatory properties and joint function. Discussion/Conclusions: The Kunitz SPI proteins synthesised by articular chondrocytes are members of the ITI superfamily. By analogy with other tissues in which these proteins occur we deduce that the cartilage Kunitz SPIs may be multifunctional proteins. Binding of the cartilage Kunitz SPIs to HA may protect this polymer from depolymerisation by free radical damage and may also protect other components in the cartilage surface from proteolytic degradation preserving joint function.

scholarly journals Comparative properties of three functionally different but structurally related serpin variants from horse plasma

1991 ◽  
Vol 274 (2) ◽  
pp. 465-471 ◽  
Author(s):  
J Potempa ◽  
J K Wunderlich ◽  
J Travis
Keyword(s):  
Peptide Bond ◽  
High Rate ◽  
Reactive Site ◽  
Association Rate ◽  
Elastase Inhibitor ◽  
Oxidative Inactivation ◽  
Methionine Residue ◽  
Horse Plasma ◽  
Oxidation Resistant ◽  
Association Rate Constants

Three structurally related but functionally different serpins from horse plasma were isolated and characterized. In spite of their identical N-terminal sequences, which show some similarity to that of human alpha 1-proteinase inhibitor, the reactive-centre loops of each of these proteins show extensive variation. Only inhibitor I, with a P1 methionine residue, resembles human alpha 1-PI with regard to (a) similarity of amino acid sequence in the vicinity of the reactive-site peptide bond, (b) broad inhibitory specificity, (c) sensitivity to oxidative inactivation and (d) high rate of reactivity with neutrophil elastase(s). Inhibitor II, with a P1 arginine residue, is an exclusive trypsin inhibitor, and inhibitor III is an oxidation-resistant slow-reacting elastase inhibitor with a P1 alanine residue. Comparison of association rate constants for the inhibition of horse neutrophil elastases by the three inhibitors indicates that only inhibitor I is likely to be physiologically important in the regulation of these enzymes.

Design of serine proteinase inhibitors by combinatorial chemistry using trypsin inhibitor SFTI-1 as a starting structure

2007 ◽  
Vol 13 (11) ◽  
pp. 749-755 ◽  
Author(s):  
Ewa Zabłotna ◽  
Anna Jaśkiewicz ◽  
Anna Łęgowska ◽  
Hanna Miecznikowska ◽  
Adam Lesner ◽  
...  
Keyword(s):  
Trypsin Inhibitor ◽  
Combinatorial Chemistry ◽  
Proteinase Inhibitors ◽  
Serine Proteinase ◽  
Serine Proteinase Inhibitors

scholarly journals Structural and functional characterization of elastases from horse neutrophils

1994 ◽  
Vol 300 (2) ◽  
pp. 401-406 ◽  
Author(s):  
A Dubin ◽  
J Potempa ◽  
J Travis
Keyword(s):  
Neutrophil Elastase ◽  
Human Neutrophil ◽  
Proteinase Inhibitors ◽  
Functional Characterization ◽  
Human Neutrophils ◽  
Human Neutrophil Elastase ◽  
Proteinase 3 ◽  
Reactive Site ◽  
Chronic Lung Diseases

In order better to understand the pathophysiology of the equine form of emphysema, two elastinolytic enzymes from horse neutrophils, referred to as proteinases 2A and 2B, have been extensively characterized and compared with the human neutrophil proteinases, proteinase-3 and elastase. Specificity studies using both the oxidized insulin B-chain and synthetic peptides revealed that cleavage of peptide bonds with P1 alanine or valine residues was preferred. Further characterization of the two horse elastases by N-terminal sequence and reactive-site analyses indicated that proteinases 2A and 2B have considerable sequence similarity to each other, to proteinase-3 from human neutrophils (proteinase 2A), to human neutrophil elastase (proteinase 2B) and to a lesser extent to pig pancreatic elastase. Horse and human elastases differed somewhat in their interaction with some natural protein proteinase inhibitors. For example, in contrast with its action on human neutrophil elastase, aprotinin did not inhibit either of the horse proteinases. However, the Val15, alpha-aminobutyric acid-15 (Abu15), alpha-aminovaleric acid-15 (Nva15) and Ala15 reactive-site variants of aprotinin were good inhibitors of proteinase 2B (Ki < 10(-9) M) but only weak inhibitors of proteinase 2A (Ki > 10(-7) M). In summary, despite these differences, the horse neutrophil elastases were found to resemble closely their human counterparts, thus implicating them in the pathological degradation of connective tissue in chronic lung diseases in the equine species.

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