HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 280, Issue 15, 15471-15478, April 15, 2005

This Article Full Text Full Text (PDF) All Versions of this Article: 280/15/15471    most recent M500881200v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Lu, G. Articles by Krishnaswamy, S. Search for Related Content PubMed PubMed Citation Articles by Lu, G. Articles by Krishnaswamy, S. Social Bookmarking                      What's this?

The Affinity of Protein C for the Thrombin·Thrombomodulin Complex Is Determined in a Primary Way by Active Site-dependent Interactions^*

Genmin Lu, Sotheavy Chhum, and Sriram Krishnaswamy¶

From the Joseph Stokes Research Institute, Children's Hospital of Philadelphia and the Department of Pediatrics, University of Pennsylvania, Philadelphia, Pennsylvania 19104

The interaction of thrombin (IIa) with thrombomodulin (TM) is essential for the efficient activation of protein C (PC). Interactions between PC and extended surfaces, likely contributed by TM within the IIa·TM complex, have been proposed to play a key role in PC activation. Initial velocities of PC activation at different concentrations of PC and TM could be accounted for by a model that did not require consideration of direct binding interactions between PC and TM. Reversible inhibitors directed toward the active site of IIa within the IIa·TM complex behaved as classic competitive inhibitors of both peptidyl substrate cleavage as well as PC activation. The ability of these small molecule inhibitors to block PC binding to the enzyme points to a principal role for active site-dependent substrate recognition in determining the affinity of IIa·TM for its protein substrate. Selective abrogation of active site docking by mutation of the P1 Arg in PC to Gln yielded an uncleavable derivative (PC_R15Q). PC_R15Q was a poor inhibitor (K_i 30 µM) of PC activation as well as peptidyl substrate cleavage by IIa·TM. Thus, inhibition by PC_R15Q most likely results from its ability to weakly interfere with active site function rather than by blocking extended interactions with the enzyme complex. The data suggest a primary role for active site-dependent substrate recognition in driving the affinity of the IIa·TM complex for its protein substrate. Interactions between PC and extended surfaces contributed by IIa and/or TM within the IIa·TM complex likely contribute in a secondary or minor way to protein substrate affinity.

Received for publication, January 24, 2005 , and in revised form, February 8, 2005.

^* This work was supported by National Institutes of Health Grants HL-47465 and HL-74124 (to S. K.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

^¶ To whom correspondence should be addressed: Joseph Stokes Research Institute, Children's Hospital of Philadelphia, 310A Abramson, 3615 Civic Center Blvd., Philadelphia, PA 19104. Tel.: 215-590-3346; Fax: 215-590-2320; E-mail: skrishna{at}mail.med.upenn.edu.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				M.-C. Bouton, L. Venisse, B. Richard, C. Pouzet, V. Arocas, and M. Jandrot-Perrus Protease Nexin-1 Interacts With Thrombomodulin and Modulates Its Anticoagulant Effect Circ. Res., April 27, 2007; 100(8): 1174 - 1181. [Abstract] [Full Text] [PDF]

				L. Yang, C. Manithody, and A. R. Rezaie Activation of protein C by the thrombin-thrombomodulin complex: Cooperative roles of Arg-35 of thrombin and Arg-67 of protein C PNAS, January 24, 2006; 103(4): 879 - 884. [Abstract] [Full Text] [PDF]

				D. A. Lane, H. Philippou, and J. A. Huntington Directing thrombin Blood, October 15, 2005; 106(8): 2605 - 2612. [Abstract] [Full Text] [PDF]