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

J. Biol. Chem., Vol. 282, Issue 22, 16095-16104, June 1, 2007

This Article Full Text Full Text (PDF) All Versions of this Article: 282/22/16095    most recent M610577200v1 Submit a Letter to Editor Alert me when this article is cited Alert me when eLetters are posted 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 Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Kroh, H. K. Articles by Bock, P. E. Search for Related Content PubMed PubMed Citation Articles by Kroh, H. K. Articles by Bock, P. E. Social Bookmarking                      What's this?

Expression of Allosteric Linkage between the Sodium Ion Binding Site and Exosite I of Thrombin during Prothrombin Activation^*

Heather K. Kroh¹, Guido Tans, Gerry A. F. Nicolaes, Jan Rosing, and Paul E. Bock²

From the Department of Pathology, Vanderbilt University, Nashville, Tennessee 37232 and the Department of Biochemistry, Cardiovascular Research Institute Maastricht, University Maastricht, Maastricht 6200MD, The Netherlands

The specificity of thrombin for procoagulant and anticoagulant substrates is regulated allosterically by Na⁺. Ordered cleavage of prothrombin (ProT) at Arg³²⁰ by the prothrombinase complex generates proteolytically active, meizothrombin (MzT), followed by cleavage at Arg²⁷¹ to produce thrombin and fragment 1.2. The alternative pathway of initial cleavage at Arg²⁷¹ produces the inactive zymogen form, the prethrombin 2 (Pre 2)·fragment 1.2 complex, which is cleaved subsequently at Arg³²⁰. Cleavage at Arg³²⁰ of ProT or prethrombin 1 (Pre 1) activates the catalytic site and the precursor form of exosite I (proexosite I). To determine the pathway of expression of Na⁺-(pro)exosite I linkage during ProT activation, the effects of Na⁺ on the affinity of fluorescein-labeled hirudin-(54–65) ([5F]Hir-(54–65)(SO^–₃)) for the zymogens, ProT, Pre 1, and Pre 2, and for the proteinases, MzT and MzT-desfragment 1 (MzT(–F1)) were quantitated. The zymogens showed no significant linkage between proexosite I and Na⁺, whereas cleavage at Arg³²⁰ caused the affinities of MzT and MzT(–F1) for [5F]Hir-(54–65)(SO^–₃) to be enhanced by Na⁺ 8- to 10-fold and 5- to 6-fold, respectively. MzT and MzT(–F1) showed kinetically different mechanisms of Na⁺ enhancement of chromogenic substrate hydrolysis. The results demonstrate for the first time that MzT is regulated allosterically by Na⁺. The results suggest that the distinctive procoagulant substrate specificity of MzT, in activating factor V and factor VIII on membranes, and the anticoagulant, membrane-modulated activation of protein C by MzT bound to thrombomodulin are regulated by Na⁺-induced allosteric transition. Further, the Na⁺ enhancement in MzT activity and exosite I affinity may function in directing the sequential ProT activation pathway by accelerating thrombin formation from the MzT fast form.

Received for publication, November 14, 2006 , and in revised form, April 4, 2007.

^* This work was supported in part by NHLBI, National Institutes of Health Grant HL038779 (to P. E. B.), and the Edmond Hustinx Foundation, Cardiovascular Research Institute Maastricht, Dept. of Biochemistry, University Maastricht. This work was also supported by the Dutch Organization for Scientific Research (VIDI Grant 916-046-330 to G. A. F. N.). 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.

¹ Supported by NIH Training Grant HL 07751.

² To whom correspondence should be addressed: Dept. of Pathology, Vanderbilt University School of Medicine, C3321A Medical Center North, Nashville, TN 37232-2561. Tel.: 615-343-9863; Fax: 615-322-1855; E-mail: paul.bock{at}vanderbilt.edu.

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

This article has been cited by other articles:

				P. S. Gandhi, Z. Chen, F. S. Mathews, and E. Di Cera Structural identification of the pathway of long-range communication in an allosteric enzyme PNAS, February 12, 2008; 105(6): 1832 - 1837. [Abstract] [Full Text] [PDF]