Interaction of heterotrimeric G protein Gαo with Purkinje cell protein-2 -: Evidence for a novel nucleotide exchange factor

被引:73
作者
Luo, Y
Denker, BM
机构
[1] Brigham & Womens Hosp, Dept Med, Div Renal, Boston, MA 02115 USA
[2] Harvard Univ, Sch Med, Boston, MA 02115 USA
关键词
D O I
10.1074/jbc.274.16.10685
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The heterotrimeric G protein G alpha(o) is ubiquitously expressed throughout the central nervous system, but many of its functions remain to be defined. To search for novel proteins that interact with G alpha(o) a mouse brain library was screened using the yeast two-hybrid interaction system. Pcp2 (Purkinje cell protein-2) was identified as a partner for G alpha(o) in this system. Pcp2 is expressed in cerebellar Purkinje cells and retinal bipolar neurons, two locations where G alpha(o) is also expressed. Pcp2 was first identified as a candidate gene to explain Purkinje cell degeneration in pcd mice (Nordquist, D. T., Kozak, C, A., and Orr, H. T, (1988) J, Neurosci, 8, 4780-4789), but its function remains unknown as Pcp2 knockout mice are normal (Mohn, A. R,, Feddersen, R, M,, Nguyen, M, S., and Koller, B, H, (1997) Mol. Cell. Neurosci, 9, 63-76), G alpha(o) and Pcp2 binding was confirmed in vitro using glutathione S-transferase-Pcp2 fusion proteins and in vitro translated [S-35]methionine-labeled G alpha(o). In addition, when G alpha(o) and Pcp2 were cotransfected into COS cells, G alpha(o) was detected in immunoprecipitates of Pcp2. To determine whether Pcp2 could modulate G alpha(o) function, kinetic constants k(cat) and k(off) of bovine brain G alpha(o) were determined in the presence and absence of Pcp2, Pcp2 stimulates GDP release from G alpha(o) more than 5-fold without affecting k(cat). These findings define a novel nucleotide exchange function for Pcpa and suggest that the interaction between Pcp2 and G alpha(o) is important to Purkinje cell function.
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页码:10685 / 10688
页数:4
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