Binding of platelet glycoprotein Ibbeta through the convex surface of leucine-rich repeats domain of glycoprotein IX.
BACKGROUND
The mechanism of assembly of the platelet glycoprotein (GP) Ib-IX complex from GPIbalpha, GPIbbeta and GPIX subunits is not entirely clear. In this complex, ectodomains of both GPIbbeta and GPIX subunits contain two leucine-rich repeats (LRR) and share high sequence similarity. However, they differ noticeably in stability, hampering further analysis of their interaction.


OBJECTIVE
Guided by analysis of the LRR structure, we report a well-folded Ibbeta/IX chimera and its usage in dissecting GPIX function.


RESULTS
In this chimera, three non-contiguous sequences that may constitute the putative convex surface of the GPIbbeta ectodomain are replaced by their GPIX counterparts. Like GPIbbeta but unlike GPIX ectodomain, it can secrete from transfected Chinese hamster ovary cells and fold into a stable conformation. Furthermore, replacing the ectodomain in GPIX with the Ibbeta/IX chimera, but not the GPIbbeta ectodomain, preserved its interaction with GPIbbeta as demonstrated by its native-like GPIbbeta-induced increase in surface expression and coimmunoprecipitation.


CONCLUSIONS
The putative convex surface of the LRR domain in GPIX is sufficient, in the context of full-length subunit, to mediate its association with GPIbbeta.
Mo
X
X
Center for Membrane Biology, Department of Biochemistry and Molecular Biology, The University of Texas Health Science Center at Houston, Houston, TX 77030, USA.
Nguyen
N X
NX
McEwan
P A
PA
Zheng
X
X
López
J A
JA
Emsley
J
J
Li
R
R
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D018076:DNA, Complementary / Q000378:metabolism; D005434:Flow Cytometry; D047468:Immunoprecipitation; D007930:Leucine / Q000737:chemistry*; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry
D018076:DNA, Complementary; D019038:Platelet Glycoprotein GPIb-IX Complex; D011993:Recombinant Fusion Proteins; D007930:Leucine