Shimono K, Iwamoto M, Sumi M, Kamo N
Laboratory of Biophysical Chemistry, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo, 060-0812, Japan.
J Biochem. 1998 Aug;124(2):404-9. doi: 10.1093/oxfordjournals.jbchem.a022126.
Crystallographic data reveal that Met-118 in bacteriorhodopsin (bR) contacts directly with the C9 methyl group of retinal, and Khorana et al. [J. Biol. Chem. 268, 20305-20311 (1993)] suggest that this contact may regulate the absorption maximum (lambdamax). We have replaced the amino acid (Val-108) corresponding to Met-118 of bR by methionine in pharaonis phoborhodopsin (ppR), whose lambdamax is ca. 500 nm, while those of other bacterial rhodopsins such as bR, halorhodopsin, and sensory rhodopsin are red-shifted by 60-90 nm. By flash-photolysis measurement, we could not recognize a large spectral red-shift of the V108M mutant. On the other hand, the decay of ppRM (M-intermediate) of the mutant was approximately three times as fast as that of wild-type, and an M-like intermediate (M') whose lambdamax is blue-shifted by 60 nm from that of M became appreciable. The replacement abolished the shoulder of the ppRM spectrum. From these findings, we infer that the distance between the retinal and the 108-position in ppR is relatively long, and that in the M-state this distance is shortened.
晶体学数据显示，细菌视紫红质（bR）中的甲硫氨酸-118直接与视黄醛的C9甲基接触，并且霍拉纳等人[《生物化学杂志》268, 20305 - 20311 (1993)]提出这种接触可能调节最大吸收波长（λmax）。我们已将法老嗜盐菌视紫红质（ppR）中对应于bR的甲硫氨酸-118的氨基酸（缬氨酸-108）替换为甲硫氨酸，其λmax约为500纳米，而其他细菌视紫红质如bR、嗜盐菌视紫红质和感官视紫红质的λmax红移了60 - 90纳米。通过闪光光解测量，我们未识别出V108M突变体有大的光谱红移。另一方面，该突变体的ppRM（M中间体）的衰减速度约为野生型的三倍，并且出现了一种λmax比M蓝移60纳米的类M中间体（M'）。这种替换消除了ppRM光谱的肩部。从这些发现中，我们推断ppR中视黄醛与108位之间的距离相对较长，并且在M态下这个距离会缩短。
