Bovine Protein C Inhibitor Has a Unique Reactive Site and Can Transiently Inhibit Plasmin

2000 ◽  
Vol 83 (02) ◽  
pp. 262-267 ◽  
Author(s):  
Hiroyuki Yuasa ◽  
Hitoshi Tanaka ◽  
Tatsuya Hayashi ◽  
Toshiaki Wakita ◽  
Hideaki Nakamura ◽  
...  
Keyword(s):  
Protein C ◽  
Activated Protein C ◽  
Peptide Bond ◽  
Serine Protease Inhibitor ◽  
Mature Protein ◽  
Reactive Site ◽  
Plasmin Activity ◽  
Amino Acid Sequence Homology ◽  
Hemostatic System ◽  
Protein C Inhibitor

SummaryProtein C inhibitor (PCI) regulates the anticoagulant protein C pathway by neutralizing activated protein C and thrombin-thrombomodulin complex in the human hemostatic system. In this study, we cloned a full-length bovine PCI cDNA encoding a putative 19-residue signal peptide and a 385-residue mature protein; this showed 70.6%, 70.6%, 57.5% and 59.6% amino acid sequence homology with the human, rhesus monkey, rat and mouse PCIs, respectively. Bovine PCI mRNA (2.1 kb in size) was expressed strongly in the liver, and moderately in the kidney and testis, but not in other tissues tested. Bovine PCI has a putative reactive site peptide bond, Lys-Ser, that is different from the reactive site sequence (Arg-Ser) of other species’ PCI. We found that bovine PCI transiently inhibits bovine plasmin, but not human plasmin. Western blot analysis showed that the reactive site of bovine PCI is cleaved during the course of complex formation with bovine plasmin; degraded PCI is released from the complex gradually concomitant with the recovery of plasmin activity. These findings suggest that bovine PCI plays a role not only in the protein C pathway but also in the fibrinolytic activity of bovine hemostatic system. Abbreviations: PCI, protein C inhibitor, Serpin, serine protease inhibitor, APC, activated protein C, TM, thrombomodulin.

Involvement of Lys 62[217] and Lys 63[218] of Human Anticoagulant Protein C in Heparin Stimulation of Inhibition by the Protein C Inhibitor

1999 ◽  
Vol 82 (07) ◽  
pp. 72-79 ◽  
Author(s):  
Lei Shen ◽  
Bruno Villoutreix ◽  
Björn Dahlbäck
Keyword(s):  
Rate Constants ◽  
Protein C ◽  
Double Mutant ◽  
Activated Protein C ◽  
Human Kidney ◽  
Peptide Bond ◽  
Dimensional Structure ◽  
Heparin Binding ◽  
Protein C Inhibitor ◽  
Stimulation Of

SummaryInhibition of activated protein C (APC) by protein C inhibitor (PCI) is stimulated by heparin, whereas inhibition by α1-antitrypsin (AAT) is heparin-independent. Three lysine residues located in a positively charged cluster in the serine protease domain of protein C (PC) were mutated to probe their involvement in the heparin stimulation of inhibition by PCI. These mutations were selected after analysis of the three-dimensional structure of APC and of molecular models for PCI and the APC-PCI complex. A double mutant, K62[217]N/K63[218]D, a single mutant, K86[241]S, and wild-type PC were expressed in embryonic human kidney 293 cells. Heparin stimulated the rate of inhibition of wt-APC by PCI approximately 400-fold, with second order rate constants (k 2 ) in the absence and presence of heparin of 0.72 × 103 M–1s–1 and 2.87 × 105 M–1s–1, respectively. In contrast, heparin only yielded a 52-fold stimulation of the rate of inhibition of the double mutant APC by PCI as the rate constants in the absence and presence of heparin were k 2 = 2.44 × 103 M–1s–1 and k 2 = 1.26 × 105 M–1s–1, respectively. The double mutant K62N/K63D eluted at approximately 10% lower NaCl concentration from a heparin Sepharose column than the K86S mutant or wt-APC. These data suggest K62 and K63 in APC to be part of a heparin binding site which is important for heparin-mediated stimulation of inhibition of APC by PCI. Abbreviations: APC, activated protein C; PC, protein C; PCI, protein C inhibitor, AAT, α1-antitrypsin also called α1-proteinase inhibitor, AT, antithrombin; TM, thrombomodulin. The chymotrypsinogen nomenclature for APC (1) is used in the text while the PC numbering is indicated between brackets whenever appropriate. P1, P2... and P1’, P2’.. designate inhibitor residues amino- and carboxy-terminal to the scissile peptide bond, respectively, and S1, S2.. and S1’, S2’.. the corresponding subsites of the protease (2). The antitrypsin numbering (3) for PCI is used along this article while the PCI numbering is mentioned between brackets whenever appropriate.

Characterization of a cDNA for Rhesus Monkey Protein C Inhibitor – Evidence for N-Terminal Involvement in Heparin Stimulation

1995 ◽  
Vol 74 (04) ◽  
pp. 1079-1087 ◽  
Author(s):  
Klaus-P Radtke ◽  
José A Fernández ◽  
Bruno O Villoutreix ◽  
Judith S Greengard ◽  
John H Griffin
Keyword(s):  
Rhesus Monkey ◽  
Protein C ◽  
Activated Protein C ◽  
Pcr Amplification ◽  
Amino Acid Sequences ◽  
Heparin Binding ◽  
Reactive Center ◽  
Protein C Inhibitor ◽  
Polymerase Chain

SummarycDNAs for protein C inhibitor (PCI) were cloned from human and rhesus monkey 1 liver RNAs by reverse transcription and polymerase chain reaction (PCR) amplification. Sequencing showed that rhesus monkey and human PCI cDNAs were 93% identical. Predicted amino acid sequences differed at 26 of 387 residues. Pour of these differences (T352M, N359S, R362K, L3631) were in the reactive center loop that is important for inhibitory specificity, and two were in the N-terminal helix (M8T, E13K) that is implicated in glycosaminoglycan binding. PCI in human or rhesus monkey plasma showed comparable inhibitory activity towards human activated protein C in the presence of 10 U/ml heparin. However, maximal acceleration of the inhibition of activated protein C required 5-fold lower heparin concentration for rhesus monkey than for human plasma, consistent with the interpretation that the additional positive charge (E13K) in a putative-heparin binding region increased the affinity for heparin.

scholarly journals Plasma levels of activated protein C-protein C inhibitor complex in patients with hypercoagulable states

2000 ◽  
Vol 65 (1) ◽  
pp. 35-40 ◽  
Author(s):  
Rika Watanabe ◽  
Hideo Wada ◽  
Miho Sakakura ◽  
Yoshitaka Mori ◽  
Takahiro Nakasaki ◽  
...  
Keyword(s):  
Plasma Levels ◽  
Protein C ◽  
Activated Protein C ◽  
C Protein ◽  
Inhibitor Complex ◽  
Protein C Inhibitor

scholarly journals Activated protein C–protein C inhibitor complex as a prognostic marker in sepsis

Critical Care ◽  
2008 ◽  
Vol 12 (Suppl 5) ◽  
pp. P21
Author(s):  
Lars Heslet ◽  
Rikke Hald ◽  
Camilla Recke ◽  
Kristian Bangert ◽  
Lars Uttenthal
Keyword(s):  
Prognostic Marker ◽  
Protein C ◽  
Activated Protein C ◽  
C Protein ◽  
Inhibitor Complex ◽  
Protein C Inhibitor

Basic residues in the 37-loop of activated protein C modulate inhibition by protein C inhibitor but not by α1-antitrypsin

2003 ◽  
Vol 1649 (1) ◽  
pp. 106-117 ◽  
Author(s):  
Laura N. Glasscock ◽  
Bruce Gerlitz ◽  
Scott T. Cooper ◽  
Brian W. Grinnell ◽  
Frank C. Church
Keyword(s):  
Protein C ◽  
Activated Protein C ◽  
Protein C Inhibitor

Activated Protein C-Protein C Inhibitor Complex in Peripheral Arterial Disease

2010 ◽  
Vol 24 (5) ◽  
pp. 588-595 ◽  
Author(s):  
David Blomstrand ◽  
Tilo Kölbel ◽  
Bengt Lindblad ◽  
Anders Gottsäter
Keyword(s):  
Peripheral Arterial Disease ◽  
Protein C ◽  
Activated Protein C ◽  
Arterial Disease ◽  
C Protein ◽  
Inhibitor Complex ◽  
Protein C Inhibitor ◽  
Peripheral Arterial

scholarly journals Turnover of *I-protein C inhibitor and *I-alpha 1-antitrypsin and their complexes with activated protein C

Blood ◽  
1990 ◽  
Vol 76 (11) ◽  
pp. 2290-2295 ◽  
Author(s):  
M Laurell ◽  
J Stenflo ◽  
TH Carlson
Keyword(s):  
Complex Formation ◽  
Protein C ◽  
Activated Protein C ◽  
Human Protein ◽  
Relative Contribution ◽  
Protein C Inhibitor ◽  
The Difference ◽  
Alpha 1 Antitrypsin

Abstract The rates of clearance and catabolism of human protein C inhibitor (PCI) and human alpha 1-antitrypsin (alpha 1-AT) and their complexes with human activated protein C (APC) were studied in the rabbit. The radioiodinated-free inhibitors had biologic half-lives of 23.4 and 62.1 hours, respectively, while the corresponding *I-labeled activated- protein C complexes were cleared with half-lives of 19.6 +/- 3.1 and 72.2 +/- 6.1 minutes. Complex clearances were linked to their catabolism as shown by a correlation between clearance and the appearance of free radioiodine in the plasma. Thus, the difference in the rates of catabolism would result in a fivefold greater amount of alpha 1-AT-APC complex than PCI-APC complex 1 hour after the formation of equal amounts of these in vivo. These results lead to the conclusion that the relative contribution of PCI and alpha 1-AT to the physiologic inhibition of APC cannot be determined only from the rates of the formation of these complexes in vitro, or from measurement of their levels in plasma. The APC-PCI complex is unstable as compared with the APC-alpha 1-AT complex, compounding the problem of estimating rates of complex formation from their levels in plasma.

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