Selectivity of Retinal Photoisomerization in Proteorhodopsin Is Controlled by Aspartic Acid 227

• Published in: Biochemistry (Easton) Vol. 43; no. 6; pp. 1648 - 1655
• Main Authors: Imasheva, Eleonora S, Balashov, Sergei P, Wang, Jennifer M, Dioumaev, Andrei K, Lanyi, Janos K
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 17-02-2004
• Subjects: Amino Acid Substitution - genetics; Asparagine - genetics; Aspartic Acid - chemistry; Aspartic Acid - genetics; Bacteriorhodopsins - chemistry; Bacteriorhodopsins - genetics; Bacteriorhodopsins - metabolism; Binding Sites - genetics; Darkness; Gammaproteobacteria; Hydrogen-Ion Concentration; Isomerism; Light; Retinaldehyde - chemistry; Rhodopsin - chemistry; Rhodopsin - genetics; Rhodopsin - metabolism; Rhodopsins, Microbial; Solubility; Spectroscopy, Fourier Transform Infrared; Temperature
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi0355894

Similarly to bacteriorhodopsin, proteorhodopsin that normally contains all-trans and 13-cis retinal is transformed at low pH to a species containing 9-cis retinal under continuous illumination at λ > 530 nm. This species, absorbing around 430 nm, returns thermally in tens of minutes to initial pigment and can be reconverted also with blue-light illumination. The yield of the 9-cis species is negligibly small at neutral pH but increases manyfold (>100) at acid pH with a pK a of 2.6. This indicates that protonation of acidic group(s) alters the photoreaction pathway that leads normally to all-trans → 13-cis isomerization. In the D97N mutant, in which one of the two acidic groups in the vicinity of the retinal Schiff base is not ionizable, the yield of 9-cis species at low pH shows a pH dependence similar to that in the wild-type but with a somewhat increased pK a of 3.3. In contrast to this relatively minor effect, replacement of the other acidic group, Asp227, with Asn results in a remarkable, more than 50-fold, increase in the yield of the light-induced formation of 9-cis species in the pH range 4−6. It appears that protonation of Asp227 at low pH is what causes the dramatic increase in the yield of the 9-cis species in wild-type proteorhodopsin. We conclude that the photoisomerization pathways in proteorhodopsin to 13-cis or 9-cis photoproducts are controlled by the charge state of Asp227.