Roginskaya M, Connelly S M, Kim K S, Patel D, Dumont M E
Department of Biochemistry and Biophysics, University of Rochester School of Medicine and Dentistry, PO Box 712, Rochester, NY 14642, USA.
Mol Genet Genomics. 2004 Mar;271(2):237-48. doi: 10.1007/s00438-004-0975-y. Epub 2004 Feb 7.
The sites and modes of interaction between G protein-coupled receptors and their cognate heterotrimeric G proteins remain poorly defined. The C-terminus of the Galpha subunit is the best established site of contact of G proteins with receptors, but structural analyses and crosslinking studies suggest the possibility of interactions at the N-terminus of Galpha as well. We screened for mutations in the N-terminal region of the Galpha subunit encoded by the yeast GPA1 gene that specifically affect the ability of the G protein to be activated by the yeast alpha-mating factor receptor. The screen led to identification of substitutions of glutamine or proline for Leu18 of Gpa1p that reduce the response to the pheromones alpha-factor and a-factor without affecting cellular levels of the subunit or its ability to interact with beta and gamma subunits. The mutations do not appear to affect the intrinsic ability of the G protein to be converted to the activated state. The low yield of different mutations with this phenotype indicates either that the N-terminal segment of the yeast Galpha subunit does not undergo extensive interactions with the alpha-factor receptor, or that this region can not be altered without detrimental effects upon the formation of G protein trimers.
G蛋白偶联受体与其同源异源三聚体G蛋白之间的相互作用位点和方式仍不清楚。Gα亚基的C末端是G蛋白与受体最确定的接触位点，但结构分析和交联研究表明，Gα亚基的N末端也可能存在相互作用。我们筛选了酵母GPA1基因编码的Gα亚基N末端区域的突变，这些突变特异性地影响G蛋白被酵母α-交配因子受体激活的能力。该筛选导致鉴定出Gpa1p的Leu18被谷氨酰胺或脯氨酸取代，这降低了对信息素α-因子和a-因子的反应，而不影响该亚基的细胞水平或其与β和γ亚基相互作用的能力。这些突变似乎不影响G蛋白转化为激活状态的内在能力。具有这种表型的不同突变的低发生率表明，要么酵母Gα亚基的N末端片段与α-因子受体没有广泛的相互作用，要么该区域在不损害G蛋白三聚体形成的情况下不能被改变。
