V108M MUTANT OF PHARAONIS PHOBORHODOPSIN - SUBSTITUTION CAUSED NO ABSORPTION CHANGE BUT AFFECTED ITS M-STATE

Crystallographic data reveal that Met-118 in bacteriorhodopsin (bR) co ntacts directly with the C-9 methyl group of retinal, and Khorana et a d. [J. Biol. Chem, 268, 20305-20311 (1993)] suggest that this contact may regulate the absorption maximum (lambda(max)) We have replaced the amino acid (Val-108) corresponding to Met-118 of bR by methionine in pharaonis phoborhodopsin (ppR), whose lambda(max) is ca. 500 run, whil e those of other bacterial rhodopsins such as bR, halorhodopsin, and s ensory rhodopsin are Fed-shifted by 60-90nm. By flash-photolysis measu rement, we could not recognize a large spectral red-shift of the V108M mutant. On the other hand, the decay of ppR(M) (M-intermediate) of th e mutant was approximately three times as fast as that of wild-type, a nd an M-like intermediate (M') whose lambda(max) is blue-shifted by 60 nm from that of M became appreciable, The replacement abolished the s houlder of the ppR(M) spectrum, From these findings, we infer that the distance between the retinal and the 108-position in ppR is relativel y long, and that in the M-state this distance is shortened.