Hydrophobic amino acids in the retinal-binding pocket of bacteriorhodopsin.
The hydrophobic amino acids Met-20, Val-49, Ala-53, Met-118, Gly-122, and Met-145 are believed to be among the amino acids that form the retinal-binding pocket in bacteriorhodopsin. We have now replaced the above amino acids, one at a time, and report on the effects of these replacements (M20A/E, V49A/L, A53G, M118A/E, G122C, and M145A/E) on the properties of bacteriorhodopsin. With the exception of Met-20, replacements at all of the other positions resulted in (i) altered rates of in vitro chromophore formation that ranged from 20-fold faster to 45-fold slower than wild-type bacteriorhodopsin, (ii) blue shifts in the visible spectra of up to 80 nm, and (iii) caused changes in the retinal isomer compositions of the mutant chromophores. Specific effects were also observed in the photocycles of the Met-118, Met-145, and Val-49 mutants, suggesting that these 3 amino acids have important roles in light transduction by bacteriorhodopsin. These results are discussed together with previous studies on the effects of amino acid replacements in the retinal-binding pocket of bacteriorhodopsin.
Greenhalgh
D A
DA
Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
Farrens
D L
DL
Subramaniam
S
S
Khorana
H G
HG
D000595:Amino Acid Sequence; D001436:Bacteriorhodopsins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism; D003001:Cloning, Molecular; D004134:Dimyristoylphosphatidylcholine; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D019617:Halobacterium salinarum / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008081:Liposomes; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010713:Phosphatidylcholines; D010743:Phospholipids; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012172:Retinaldehyde / Q000378:metabolism*; D013053:Spectrophotometry; D013329:Structure-Activity Relationship
D008081:Liposomes; D010713:Phosphatidylcholines; D010743:Phospholipids; D011994:Recombinant Proteins; D001436:Bacteriorhodopsins; C024990:asolectin; D012172:Retinaldehyde; D004134:Dimyristoylphosphatidylcholine