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Titolo:
GLU(192)-]GLN SUBSTITUTION IN THROMBIN YIELDS AN ENZYME THAT IS EFFECTIVELY INHIBITED BY BOVINE PANCREATIC TRYPSIN-INHIBITOR AND TISSUE FACTOR PATHWAY INHIBITOR
Autore:
GUINTO ER; YE J; LEBONNIEC BF; ESMON CT;
Indirizzi:
OKLAHOMA MED RES FDN,HOWARD HUGHES MED INST,825 NE 13TH ST OKLAHOMA CITY OK 73104 OKLAHOMA MED RES FDN,HOWARD HUGHES MED INST OKLAHOMA CITY OK 73104 UNIV OKLAHOMA,HLTH SCI CTR,DEPT PATHOL OKLAHOMA CITY OK 73104 UNIV OKLAHOMA,HLTH SCI CTR,DEPT BIOCHEM & MOLEC BIOL OKLAHOMA CITY OK73104 OKLAHOMA MED RES FDN,CARDIOVASC BIOL RES PROGRAM OKLAHOMA CITY OK 73104 UNIV CAMBRIDGE,CTR MRC,DEPT HEMATOL CAMBRIDGE CB2 2QH ENGLAND
Titolo Testata:
The Journal of biological chemistry
fascicolo: 28, volume: 269, anno: 1994,
pagine: 18395 - 18400
SICI:
0021-9258(1994)269:28<18395:GSITYA>2.0.ZU;2-J
Fonte:
ISI
Lingua:
ENG
Soggetto:
PRO-ARG CHLOROMETHYLKETONE; RAY CRYSTAL-STRUCTURE; HUMAN ALPHA-THROMBIN; PROTEIN-C; ACTIVE-SITE; KUNITZ INHIBITORS; ANTITHROMBIN-III; TRP INSERTION; THROMBOMODULIN; COAGULATION;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
29
Recensione:
Indirizzi per estratti:
Citazione:
E.R. Guinto et al., "GLU(192)-]GLN SUBSTITUTION IN THROMBIN YIELDS AN ENZYME THAT IS EFFECTIVELY INHIBITED BY BOVINE PANCREATIC TRYPSIN-INHIBITOR AND TISSUE FACTOR PATHWAY INHIBITOR", The Journal of biological chemistry, 269(28), 1994, pp. 18395-18400

Abstract

Modeling studies have ascribed the remarkable resistance of thrombin to inhibition by the Kunitz type inhibitors, bovine pancreatic trypsininhibitor (BPTI), and tissue factor pathway inhibitor (TFPI), to steric inhibition by the 60-loop insertion, especially Trp(60D) (in the chymotrypsin numbering system). Indeed, deletion of Pro(60B) Pro(60C), and Trp(60D) from this loop (des-PPW) enhances BPTI inhibition (K-i = 16 nM) (Le Bonniec, B. F., Guinto, E. R., MacGillivray, R. T. A., Stone, S. R., and Esmon, C. T. (1993) J. Biol. Chem. 268, 19055-19061). Activated protein C, however, lacks an equivalent insertion loop but is nevertheless resistant to inhibition by these Kunitz inhibitors. A unique feature of thrombin and activated protein C is the presence of Glu at position 192. Substitution of Glu(192) with Gin in activated protein C dramatically enhances inhibition by BPTI and TFPI (Rezaie, A and Esmon, C. T. (1993) J. Biol. Chem. 268, 19943-19948). We now demonstrate that thrombin E192Q (the Glu(192) --> Gln mutant) is inhibited by BPTI (K-i = 24 nM) or TFPI (K-i = 14 nM) much more effectively than wildtype thrombin (K-i > 1 mu M for both inhibitors). A thrombin mutant having both the des-PPW deletion and E192Q substitution binds BPTI (K-i= 35 pM) and TFPI (K-i = 25 pM) even tighter. BPTI can displace dansylarginine N-(-3-ethyl-1,5-pentanediyl)-amide from the active site of thrombin E192Q (K-i = 19 nM), indicating that BPTI interacts directly with the S1 binding site in thrombin. The E192Q mutation and PPW deletion contribute comparably and additively to the binding energy of thrombin with the Kunitz inhibitors. We suggest that access to the active center of thrombin is less restricted than predicted from previous studies.

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Documento generato il 02/07/20 alle ore 22:32:37