Kinetic Studies on the Interaction of α<sub><b>1</b></sub>-Proteinase Inhibitor (Pittsburgh) with Trypsin-Like Serine Proteinases

Travis, James; Matheson, Nancy; George, Peter M.; Carrell, Robin W.

doi:10.1515/bchm3.1986.367.2.853

Cited by 39 publications

(23 citation statements)

References 13 publications

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“…These proteases were both inhibited by the variant protein purified from our patient's plasma, with second-order constant rates similar to those observed for the first a1-AT Pittsburgh variant (2).…”

Section: Discussionsupporting

confidence: 71%

“…This mutant, designated a,-AT Pittsburgh, or recombinant Arg 358 a,-AT, was shown to inhibit other clotting proteases (2)(3)(4), thus displaying strong functional analogy with the physiologic clotting inhibitor antithrombin III (AT III), whose reactive site consists of an Arg-Ser bond. This mutation provided an explanation for the patient's recurrent and ultimately fatal hemorrhagic diathesis (1,5), and confirmed the critical role played by a single amino acid at the reactive site of serineprotease inhibitors (serpins).…”

Section: Introductionmentioning

confidence: 99%

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Met 358 to Arg mutation of alpha 1-antitrypsin associated with protein C deficiency in a patient with mild bleeding tendency.

Vidaud¹,

Emmerich²,

Alhenc‐Gelas³

et al. 1992

J. Clin. Invest.

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The molecular defect responsible for a dramatic prolongation of all standard clotting tests discovered in a 15-yr-old boy has been identified. Initial investigations revealed the presence of an activated Factor X (Factor Xa) and thrombin inhibitor which copurified with alpha1-antitrypsin (a,-AT), thereby suggesting the occurrence of an a,-AT variant similar to a,-AT Pittsburgh.This was confirmed by dot-blot analysis and direct sequencing after amplification by the polymerase chain reaction. A G to T transition at nucleotide 10038 results in the substitution of Met to an Arg, converting a,-AT into an Arg-Ser protease inhibitor (serpin) that inhibited thrombin and Factor Xa more effectively than antithrombin III. Surprisingly, there was no bleeding history in the proband. The common mutation Z, which may explain a reduced expression of the allele bearing the Arg 358 Met mutation, was not observed in the propositus' DNA. To exclude the presence of another mutation, the coding regions and intron/exon junctions were sequenced. No other mutation was found. Recently, the patient experienced his first hemorrhagic episode at the age of 17. The level ofthe abnormal inhibitor had increased twofold 2 mo before. The large decrease in protein C concentration may account for the mild bleeding tendency in this case, despite the presence of the a,-AT Pittsburgh mutation. An abnormal protein C pattern was observed in patient's plasma, suggesting that the circulating deficiency might be due to a deleterious effect of the abnormal inhibitor on both intracellular processing and catabolism of protein C. (J. Clin.

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Section: Discussionsupporting

confidence: 71%

Section: Introductionmentioning

confidence: 99%

Met 358 to Arg mutation of alpha 1-antitrypsin associated with protein C deficiency in a patient with mild bleeding tendency.

Vidaud¹,

Emmerich²,

Alhenc‐Gelas³

et al. 1992

J. Clin. Invest.

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“…Moreover, it gained inhibitory activity against plasmin. Therefore, mutation of Met-340 into an arginine residue changed our "␣1-antitrypsin like" to an "antithrombin-like" protein as observed for Pittsburgh factor ␣1-antitrypsin (47,48). We also introduced two point mutations (A332P and L339A) in the hinge region (position P9) and in the RCL domain (position P2), respectively, predicted to invalidate the protein.…”

Section: Discussionmentioning

confidence: 87%

Anti-hemostatic Effects of a Serpin from the Saliva of the Tick Ixodes ricinus

Prévôt

Adam

Boudjeltia

et al. 2006

Journal of Biological Chemistry

109

130

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Serpins (serine protease inhibitors) are a large family of structurally related proteins found in a wide variety of organisms, including hematophagous arthropods. Protein analyses revealed that Iris, previously described as an immunomodulator secreted in the tick saliva, is related to the leukocyte elastase inhibitor and possesses serpin motifs, including the reactive center loop (RCL), which is involved in the interaction between serpins and serine proteases. Only serine proteases were inhibited by purified recombinant Iris (rIris), whereas mutants L339A and A332P were found devoid of any protease inhibitory activity. The highest K a was observed with human leukocyteelastase, suggesting that elastase-like proteases are the natural targets of Iris. In addition, mutation M340R completely changed both Iris substrate specificity and affinity. This likely identified Met-340 as amino acid P1 in the RCL. The effects of rIris and its mutants were also tested on primary hemostasis, blood clotting, and fibrinolysis. rIris increased platelet adhesion, the contact phase-activated pathway of coagulation, and fibrinolysis times in a dose-dependent manner, whereas rIris mutant L339A affected only platelet adhesion. Taken together, these results indicate that Iris disrupts coagulation and fibrinolysis via the anti-proteolytic RCL domain. One or more other domains could be responsible for primary hemostasis inhibition. To our knowledge, this is the first ectoparasite serpin that interferes with both hemostasis and the immune response.Ticks are blood-sucking arthropods that infest a large variety of vertebrate hosts (mammals, birds, reptiles, and amphibians) in many parts of the world (1). To complete their blood meal, blood-sucking arthropods express a wide range of anti-hemostatic molecules in their saliva, including vasodilators, inhibitors of the platelet aggregation, and anti-coagulants (2). Tick saliva and salivary gland extracts are also known to modulate the host's defense mechanisms (3-6). Both anti-hemostatic and immunosuppressive compounds were identified, isolated, and characterized from soft and hard ticks. These compounds include histamine-binding proteins, tissue factor pathway inhibitor-like proteins, anti-thrombin-like proteins, and anticomplement factors (7-16).These last years, several laboratories reported the construction and screening of cDNA libraries from tick salivary glands. Thus, Das et al. (17) found 14 Ixodes scapularis immunodominant antigens, whereas Leboulle et al. (18) identified 27 mRNA, the expression of which is specifically induced or up-regulated during the Ixodes ricinus blood meal. Finally, Ribeiro and coworkers explored the sialome of the tick I. scapularis (19,20) and uncovered a large variety of putative bioactive agents. These studies all identified some serine protease inhibitors, containing serpin, kunitz, kazal, or ␣-macroglobulin motifs (21).To date, ϳ500 serpins have been identified in a large variety of species, including animals, viruses, and plants. On average, serpins are 35...

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“…This is impressively shown for the Met ~ Arg exchange in t~rproteinase inhibitor which decreases the/Con rate constant by 4 orders of magnitude for the interaction with elastase but increases this value 4 orders of magnitude for the interaction with thrombin [20]. As inhibitors react in a substrate-like manner with their proteinases one of the basic amino acid residues was thought to be the reactive site of hirudin.…”

Section: Discussionmentioning

confidence: 95%

Interaction of site specific hirudin variants with α‐thrombin

Dodt

Köhler

Baici³

1988

FEBS Letters

105

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The kinetics of complex formation between recombinant hirudin or recombinant hirudin mutants with thrombin were analyzed. In order to elucidate the inhibitor's reactive site peptide bond predetermined amino acid substitutions were introduced at positions of basic amino acid residues by means of site-directed mutagenesis of a hirudin gene. In comparison to recombinant hirudin (Ki = 19 pM) only those mutant inhibitors which were modified at amino acid position Lys 47 showed a higher K~ value for their complexes with thrombin. The observed effects are mainly due to increased kofr rate constants.

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Kinetic Studies on the Interaction of α₁-Proteinase Inhibitor (Pittsburgh) with Trypsin-Like Serine Proteinases

Cited by 39 publications

References 13 publications

Met 358 to Arg mutation of alpha 1-antitrypsin associated with protein C deficiency in a patient with mild bleeding tendency.

Met 358 to Arg mutation of alpha 1-antitrypsin associated with protein C deficiency in a patient with mild bleeding tendency.

Anti-hemostatic Effects of a Serpin from the Saliva of the Tick Ixodes ricinus

Interaction of site specific hirudin variants with α‐thrombin

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Kinetic Studies on the Interaction of α1-Proteinase Inhibitor (Pittsburgh) with Trypsin-Like Serine Proteinases

Cited by 39 publications

References 13 publications

Met 358 to Arg mutation of alpha 1-antitrypsin associated with protein C deficiency in a patient with mild bleeding tendency.

Met 358 to Arg mutation of alpha 1-antitrypsin associated with protein C deficiency in a patient with mild bleeding tendency.

Anti-hemostatic Effects of a Serpin from the Saliva of the Tick Ixodes ricinus

Interaction of site specific hirudin variants with α‐thrombin

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Kinetic Studies on the Interaction of α₁-Proteinase Inhibitor (Pittsburgh) with Trypsin-Like Serine Proteinases