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

J. Biol. Chem., Vol. 282, Issue 32, 23517-23524, August 10, 2007

This Article Full Text Full Text (PDF) Supplemental Data All Versions of this Article: 282/32/23517    most recent M701826200v1 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 Fontayne, A. Articles by Deckmyn, H. Search for Related Content PubMed PubMed Citation Articles by Fontayne, A. Articles by Deckmyn, H. Social Bookmarking                      What's this?

Paratope and Epitope Mapping of the Antithrombotic Antibody 6B4 in Complex with Platelet Glycoprotein Ib^*

Alexandre Fontayne¹, Bauke De Maeyer², Marc De Maeyer^¶, Mayo Yamashita^||, Tadashi Matsushita^**, and Hans Deckmyn^¶3

From the Laboratory for Thrombosis Research, IRC, the Laboratory of Biomolecular Modeling, Department of Chemistry, and ^¶BioMacS, KU Leuven Campus Kortrijk, E. Sabbelaan 53, B-8500 Kortrijk, Belgium, the ^||Department of Medical Technology, Nagoya University School of Health Sciences, Nagoya 466-8560, Japan, and the ^**Department of Hematology, Nagoya University Graduate School of Medicine, Nagoya 466-8560, Japan

The monoclonal antibody 6B4 has a potent antithrombotic effect in nonhuman primates by binding to the flexible loop, also known as the -switch region (amino acids 230-242), of glycoprotein Ib (GPIb). This interaction blocks, in high shear stress conditions, the specific interaction between GPIb and von Willebrand factor suppressing platelet deposition to the damaged vessel wall, a key event in the pathogenesis of arterial thrombosis. To understand the interactions between this antibody and its antigen at the amino acid level, we here report the identification of the paratope and epitope in 6B4 and GPIb, respectively, by using computer modeling and site-directed mutagenesis. The docking programs ZDOCK (rigid body docking) and HADDOCK (flexible docking) were used to model the interaction of 6B4 with GPIb and to delineate the respective paratope and epitope. 6B4 and GPIb mutants were constructed and assayed for their capacity to bind GPIb and 6B4, respectively. From these data, it is found that the paratope of 6B4 is mainly formed by five residues: Tyr^27D, Lys^27E, Asp²⁸, and Glu⁹³ located in light chain CDR1 and -3, respectively, and Tyr^100C of the heavy chain CDR3. These residues form a valley, where the GPIb flexible loop can bind via residues Asp²³⁵ and Lys²³⁷. The experimental results were finally used to build a more accurate docking model. Taken together, this information provides guidelines for the design of new derivatized lead compounds with antithrombotic properties.

Received for publication, March 2, 2007 , and in revised form, May 30, 2007.

^* This work was supported by Instituut voor de Aanmoediging van Innovatie door Wetenschap en Technologie in Vlaanderen Grant IWT 020473 and by the Sankyo Foundation of Life Science. 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.

The on-line version of this article (available at http://www.jbc.org) contains supplemental Table S1.

¹ An EU-Research Training Network (HPRN-CT-2002-00253) postdoctoral fellow.

² A bursary of the IWT.

³ To whom correspondence should be addressed. Tel.: 32-56-246422; Fax: 32-56-246997; E-mail: Hans.Deckmyn{at}kuleuven-kortrijk.be.

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

This article has been cited by other articles:

				R. Barderas, J. Desmet, P. Timmerman, R. Meloen, and J. I. Casal Affinity maturation of antibodies assisted by in silico modeling PNAS, July 1, 2008; 105(26): 9029 - 9034. [Abstract] [Full Text] [PDF]