Lee C H, Katz A, Simon M I
Division of Biology, California Institute of Technology, Pasadena 91125.
Mol Pharmacol. 1995 Feb;47(2):218-23.
The C5a chemoattractant factor receptor, when expressed in COS-7 cells, can stimulate phosphoinositide-specific phospholipase C activity through the activation of the G16 isoform of the heterotrimeric G protein, but not through the G11 isoform. To identify the regions of the G alpha 16 subunit protein that are responsible for its activation by the C5a receptor, a series of chimeras between G alpha 16 and G alpha 11 were constructed and tested for their ability to be activated by the C5a receptor. Co-transfection experiments with chimeras in which the carboxyl-terminal regions of G alpha 11 were replaced with the corresponding regions of G alpha 16 demonstrated that changes in the carboxyl terminus, e.g., replacement of 134 amino acids, were not sufficient to confer receptor specificity. An additional segment encompassing residues 220-240 of G alpha 16 was required to confer C5a-induced activation. Testing of a reciprocal series of chimeras composed of G alpha 16 sequences at the amino terminus and G alpha 11 sequences at the carboxyl terminus revealed that certain sequences extending from the amino terminus to amino acid 209 of G alpha 16 were sufficient to endow the chimera with much of the specificity for C5a-induced activation. These results suggest that receptor specificity may involve specific conformations of the G protein stabilized by concerted interactions of multiple amino acid sequences distributed throughout the G alpha protein.
C5a趋化因子受体在COS - 7细胞中表达时，可通过激活异源三聚体G蛋白的G16亚型来刺激磷酸肌醇特异性磷脂酶C的活性，但不能通过G11亚型。为了确定Gα16亚基蛋白中负责被C5a受体激活的区域，构建了一系列Gα16和Gα11之间的嵌合体，并测试它们被C5a受体激活的能力。用Gα11的羧基末端区域被Gα16的相应区域取代的嵌合体进行共转染实验表明，羧基末端的变化，例如134个氨基酸的替换，不足以赋予受体特异性。还需要一个包含Gα16第220 - 240位残基的额外片段来赋予C5a诱导的激活作用。对一系列由Gα16氨基末端序列和Gα11羧基末端序列组成的反向嵌合体的测试表明，从Gα16氨基末端延伸至第209位氨基酸的某些序列足以使嵌合体具有对C5a诱导激活的大部分特异性。这些结果表明，受体特异性可能涉及通过分布在整个Gα蛋白中的多个氨基酸序列的协同相互作用而稳定的G蛋白的特定构象。
