Ha J H, Dhanasekaran N, Koh H C, Lee C H
Department of Pharmacology and Institute of Biomedical Sciences, College of Medicine Hanyang University, Seoul, 133-791, Korea.
Biochem Biophys Res Commun. 2000 Nov 19;278(2):426-31. doi: 10.1006/bbrc.2000.3817.
Our previous study suggested that the region encompassing residues 220-240 on G(alpha16) is important in coupling with C5a receptor (Lee et al. (1995) Mol. Pharmacol. 47, 218-223). When aligned sequences are compared in the residue 220-240 segment of G(alpha16), there is a block of eight amino acids extending from residue 227 to residue 234 (227-Ile-Ala-Leu-Ile-Tyr-Leu-Ala-Ser-234) in G(alpha16) that is replaced by a heterologous block extending from amino acid residue 224 to residue 231 (224-Thr-Ser-Ile-Met-Phe-Leu-Val-Ala-231) in G(alpha11). In order to identify the specific amino acid residue necessary for coupling to C5a receptor within the extension of eight amino acids in G(alpha16), a series of chimeric G(alpha11)/G(alpha16) cDNA constructs and mutant G(alpha16) cDNAs were expressed. Then the ability of chimeras and mutant proteins to mediate C5a-induced release of inositol phosphate in transfected Cos-7 cells was tested. The results show that single amino acid Ala(228) is responsible for conferring about 40-50% of the activity of G(alpha16) induced by C5a receptor stimulation.
我们之前的研究表明，G(α16)上包含220 - 240位残基的区域在与C5a受体偶联中起重要作用（Lee等人，(1995)《分子药理学》47卷，218 - 223页）。当比较G(α16)中220 - 240位残基片段的比对序列时，在G(α16)中有一个由8个氨基酸组成的区域，从227位残基延伸至234位残基（227 - 异亮氨酸 - 丙氨酸 - 亮氨酸 - 异亮氨酸 - 酪氨酸 - 亮氨酸 - 丙氨酸 - 丝氨酸 - 234），而在G(α11)中该区域被一个从224位氨基酸残基延伸至231位残基（224 - 苏氨酸 - 丝氨酸 - 异亮氨酸 - 甲硫氨酸 - 苯丙氨酸 - 亮氨酸 - 缬氨酸 - 丙氨酸 - 231）的异源区域所取代。为了确定G(α16)中8个氨基酸延伸区域内与C5a受体偶联所需的特定氨基酸残基，我们表达了一系列嵌合的G(α11)/G(α16) cDNA构建体和突变的G(α16) cDNA。然后测试了嵌合体和突变蛋白在转染的Cos - 7细胞中介导C5a诱导的肌醇磷酸释放的能力。结果表明，单个氨基酸丙氨酸（228）赋予了约40 - 50%的由C5a受体刺激诱导的G(α16)活性。
