NDLI: Specific cysteines in beta3 are involved in disulfide bond exchange-dependent and -independent activation of alphaIIbbeta3.

Content Provider	World Health Organization (WHO)-Global Index Medicus
Author	Seligsohn, Uri Landau, Meytal Lahav, Judith Mor-cohen, Ronit Rosenberg, Nurit
Description	Author Affiliation: Mor-Cohen R ( Amalia Biron Research Institute of Thrombosis and Hemostasis, Chaim Sheba Medical Center, Tel-Hashomer 52621.)
Abstract	Disulfide bond exchange among cysteine residues in epidermal growth factor (EGF)-like domains of beta3 was suggested to be involved in activation of alphaIIbbeta3. To investigate the role of specific beta3 cysteines in alphaIIbbeta3 expression and activation, we expressed in baby hamster kidney cells normal alphaIIb with normal beta3 or beta3 with single or double cysteine substitutions of nine disulfide bonds in EGF-3, EGF-4, and beta-tail domains and assessed alphaIIbbeta3 surface expression and activation state by flow cytometry using P2 or PAC-1 antibodies, respectively. Most mutants displayed reduced surface expression of alphaIIbbeta3. Disruptions of disulfide bonds in EGF-3 yielded constitutively active alphaIIbbeta3, implying that these bonds stabilize the inactive alphaIIbbeta3 conformer. Mutants of the Cys-567-Cys-581 bond in EGF-4 were inactive even after exposure to alphaIIbbeta3-activating antibodies, indicating that this bond is necessary for activating alphaIIbbeta3. Disrupting Cys-560-Cys-583 in the EGF-3/EGF-4 or Cys-608-Cys-655 in beta-tail domain resulted in alphaIIbbeta3 activation only when Cys-560 or Cys-655 of each pair was mutated but not when their partners (Cys-583, Cys-608) or both cysteines were mutated, suggesting that free sulfhydryls of Cys-583 and Cys-608 participate in alphaIIbbeta3 activation by a disulfide bond exchange-dependent mechanism. The free sulfhydryl blocker dithiobisnitrobenzoic acid inhibited 70% of anti-LIBS6 antibody-induced activation of wild-type alphaIIbbeta3 and had a smaller effect on mutants, implicating disulfide bond exchange-dependent and -independent mechanisms in alphaIIbbeta3 activation. These data suggest that different disulfide bonds in beta3 EGF and beta-tail domains play variable structural and regulatory roles in alphaIIbbeta3.
ISSN	00219258
e-ISSN	1083351X
Journal	Journal of Biological Chemistry
Issue Number	28
Volume Number	283
Language	English
Publisher	American Society for Biochemistry and Molecular Biology (United States)
Publisher Date	2008-07-11
Publisher Place	United States
Access Restriction	Open
Subject Keyword	Cysteine Metabolism Disulfides Platelet Glycoprotein GPIIb-IIIa Complex Amino Acid Substitution Animals Antibodies, Monoclonal Chemistry Cell Line Cricetinae Genetics Protein Structure, Quaternary Protein Structure, Tertiary Biochemistry Molecular Biology
Content Type	Text
Resource Type	Article
Subject	Cell Biology Biochemistry Molecular Biology

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