Coupling between the retinal thermal isomerization and the Glu194 residue of bacteriorhodopsin.
Glu194 is a residue located at the end of F helix on the extracellular side of the light-induced proton pump bacteriorhodopsin (BR). Currently, it is well recognized that Glu194 and Glu204 residues, along with water clusters, constitute the proton release group of BR. Here we report that the replacement of Glu194 for Gln affects not only the photocycle of the protein but also has tremendous effect on the all-trans to 13-cis thermal isomerization. We studied the pH dependence of the dark adaptation of the E194Q mutant and performed HPLC analysis of the isomer compositions of the light- and partially dark-adapted states of the mutant at several pH values. Our data confirmed that E194Q exhibits extremely slow dark adaptation over a wide range of pH. HPLC data showed that a significantly larger concentration of all-trans isomer was present in the samples of the E194Q mutant even after prolonged dark adaptation. After 14 days in the dark the 13-cis to all-trans ratio was 1:3 in the mutant, compared to 2:1 in the wild type. These data clearly indicate the involvement of Glu194 in control of the rate of all-trans to 13-cis thermal isomerization.
Lazarova
Tzvetana
T
Departament de Bioquímica i de Biologia Molecular, Universitat Autònoma de Barcelona, Bellaterra, Spain. tzvetana.lazarova@uab.cat
Querol
Enric
E
Padrós
Esteve
E
D000220:Adaptation, Biological; D001436:Bacteriorhodopsins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D018698:Glutamic Acid / Q000235:genetics / Q000378:metabolism*; D019617:Halobacterium salinarum / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000528:radiation effects; D006863:Hydrogen-Ion Concentration; D007536:Isomerism; D007700:Kinetics; D008027:Light; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011984:Sensory Receptor Cells / Q000737:chemistry* / Q000378:metabolism*; D013053:Spectrophotometry; D013696:Temperature
D020033:Protein Isoforms; D018698:Glutamic Acid; D001436:Bacteriorhodopsins