天冬氨酸替代影响细菌视紫红质的质子转运。
Aspartic acid substitutions affect proton translocation by bacteriorhodopsin.
作者信息
Mogi T, Stern L J, Marti T, Chao B H, Khorana H G
机构信息
Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
出版信息
Proc Natl Acad Sci U S A. 1988 Jun;85(12):4148-52. doi: 10.1073/pnas.85.12.4148.
Abstract
We have substituted each of the aspartic acid residues in bacteriorhodopsin to determine their possible role in proton translocation by this protein. The aspartic acid residues were replaced by asparagines; in addition, Asp-85, -96, -115, and -112 were changed to glutamic acid and Asp-212 was also replaced by alanine. The mutant bacteriorhodopsin genes were expressed in Escherichia coli and the proteins were purified. The mutant proteins all regenerated bacteriorhodopsin-like chromophores when treated with a detergent-phospholipid mixture and retinal. However, the rates of regeneration of the chromophores and their lambda max varied widely. No support was obtained for the external point charge model for the opsin shift. The Asp-85----Asn mutant showed not detectable proton pumping, the Asp-96----Asn and Asp-212----Glu mutants showed less than 10% and the Asp-115----Glu mutant showed approximately equal to 30% of the normal proton pumping. The implications of these findings for possible mechanisms of proton translocation by bacteriorhodopsin are discussed.
摘要
我们已将细菌视紫红质中的每个天冬氨酸残基进行替换，以确定它们在该蛋白质质子转运过程中可能发挥的作用。天冬氨酸残基被天冬酰胺取代；此外，天冬氨酸-85、-96、-115和-112被替换为谷氨酸，天冬氨酸-212也被丙氨酸取代。突变的细菌视紫红质基因在大肠杆菌中表达，蛋白质被纯化。当用去污剂-磷脂混合物和视黄醛处理时，突变蛋白质均再生出类似细菌视紫红质的发色团。然而，发色团的再生速率及其最大吸收波长差异很大。视蛋白位移的外部点电荷模型未得到支持。天冬氨酸-85→天冬酰胺突变体未检测到质子泵浦，天冬氨酸-96→天冬酰胺和天冬氨酸-212→谷氨酸突变体显示不到正常质子泵浦的10%，天冬氨酸-115→谷氨酸突变体显示约为正常质子泵浦的30%。讨论了这些发现对细菌视紫红质质子转运可能机制的影响。
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Proc Natl Acad Sci U S A. 1988 Jun;85(12):4148-52. doi: 10.1073/pnas.85.12.4148.
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