Lee C H, Park D, Wu D, Rhee S G, Simon M I
Division of Biology, California Institute of Technology, Pasadena 91125.
J Biol Chem. 1992 Aug 15;267(23):16044-7.
The relative specificities of members of the G alpha q family of GTP-binding proteins were tested for their ability to activate different phosphoinositide-specific phospholipase C (PI-PLC) beta isozymes. Cos-7 cells were transfected with cDNA corresponding to G alpha q, G alpha 11, G alpha 14, and G alpha 16. Most of the recombinant protein was bound to the cell membrane and these membranes were washed to elute endogenous PI-PLC activity. The membrane preparation was reconstituted with purified preparations of the PI-PLC beta isozymes and guanosine 5'-O-thiotriphosphate (GTP gamma S)-stimulated enzyme activity was measured. All four proteins of the G alpha q family were found to stimulate PI-PLC beta 1, with G alpha q and G alpha 11 being most efficient. On the other hand, G alpha 16 was found to most effectively activate PI-PLC beta 2, while G alpha q, G alpha 11, and G alpha 14 showed less stimulation. Specific anti- G alpha 16 antibody blocked the stimulation of both PI-PLC beta 1 and PI-PLC beta 2 in the enriched membrane fraction. We conclude that there is specificity in the interaction of different members of the Gq family with different PI-PLC beta effectors. This specificity may be important in generating tissue- or receptor-specific responses in vivo.
对GTP结合蛋白Gαq家族成员激活不同磷脂酰肌醇特异性磷脂酶C(PI-PLC)β同工酶的能力进行了相对特异性测试。用对应于Gαq、Gα11、Gα14和Gα16的cDNA转染Cos-7细胞。大多数重组蛋白与细胞膜结合,这些细胞膜经过洗涤以洗脱内源性PI-PLC活性。用PI-PLCβ同工酶的纯化制剂对膜制剂进行重组,并测定鸟苷5'-O-硫代三磷酸(GTPγS)刺激的酶活性。发现Gαq家族的所有四种蛋白均能刺激PI-PLCβ1,其中Gαq和Gα11最为有效。另一方面,发现Gα16最有效地激活PI-PLCβ2,而Gαq、Gα11和Gα14的刺激作用较小。特异性抗Gα16抗体阻断了富集膜组分中PI-PLCβ1和PI-PLCβ2的刺激。我们得出结论,Gq家族的不同成员与不同的PI-PLCβ效应器之间的相互作用存在特异性。这种特异性在体内产生组织或受体特异性反应中可能很重要。