Sosa-Peinado A, Mueller D M
Department of Biological Chemistry, The Chicago Medical School, North Chicago, Illinois 60064, USA.
Arch Biochem Biophys. 1997 Jan 1;337(1):27-33. doi: 10.1006/abbi.1996.9761.
Residue beta-V198 of the yeast mitchondrial F1-ATPase abuts the P-loop motif and the side chain is within 3.8 A of the nucleotide as shown in the crystal structure of the bovine ATPase [J. P. Abrahams, A. G. W. Leslie, R. Lutter, and J. E. Walker (1984) Nature 370,621-628]. This study has made and analyzed 17 replacements of V198 to understand the importance of the side chain in the nucleotide binding site. In addition, a suppressor of V198S, beta-L390F, was studied in the presence of various replacements at position 198. In vivo and in vitro analyses indicate that the Val side chain is critical for forming a stable and active enzyme. Biochemical analysis of mitochondria isolated from the mutant strains indicates that amino acids with hydrophobic side chains are the most effective replacements. In addition, size is important, but a large side chain can be largely compensated for until the size reaches that of the Phe and Trp. A methyl group is the minimal side chain necessary for function, as the beta-subunit is not stable in vivo with Gly at position 198. These results indicate that V198 forms critical hydrophobic interactions with the adenine ring of the nucleotide.
酵母线粒体F1 - ATP酶的β - V198残基紧邻P环基序，且如牛ATP酶的晶体结构所示[J. P. 亚伯拉罕斯、A. G. W. 莱斯利、R. 卢特和J. E. 沃克（1984年）《自然》370, 621 - 628]，其侧链距核苷酸在3.8埃以内。本研究对V198进行了17种替换并加以分析，以了解侧链在核苷酸结合位点中的重要性。此外，还在198位存在各种替换的情况下研究了V198S的抑制子β - L390F。体内和体外分析表明，缬氨酸侧链对于形成稳定且有活性的酶至关重要。对从突变菌株分离的线粒体进行的生化分析表明，具有疏水侧链的氨基酸是最有效的替换。此外，大小很重要，但大的侧链在尺寸达到苯丙氨酸和色氨酸的尺寸之前可在很大程度上得到补偿。甲基是功能所需的最小侧链，因为在198位为甘氨酸时β亚基在体内不稳定。这些结果表明，V198与核苷酸的腺嘌呤环形成关键的疏水相互作用。
