title: A pharaonis phoborhodopsin mutant with the same retinal binding site residues as in bacteriorhodopsin.
abstract: pharaonis phoborhodopsin (ppR, also called pharaonis sensory rhodopsin II, psR-II) is a photoreceptor for negative phototaxis in Natronobacterium pharaonis. ppR has a blue-shifted absorption maximum (500 nm) relative those of other archaeal rhodopsins such as the proton-pump bacteriorhodopsin (BR; 570 nm). Among the 25 amino acids that are within 5 A of the retinal chromophore, 10 are different in BR and ppR, and they are presumed to be crucial in determining the color of their chromophores. However, the spectral red shift in a multiple mutant of ppR, in which the retinal binding site was made similar to that of BR (BR/ppR), was smaller than 40% (lambda(max) = 524 nm) than expected. In the paper presented here, we report on low-temperature Fourier transform infrared (FTIR) spectroscopy of BR/ppR, and compare the infrared spectral changes before and after photoisomerization with those for ppR and BR. The C[bond]C stretch and hydrogen out-of-plane (HOOP) vibrations of BR/ppR were similar to those of BR, suggesting that the surrounding protein moiety of BR/ppR becomes like BR. However, BR/ppR exhibited a unique IR band regarding the hydrogen bond of the protonated Schiff base. It has been known that ppR has a stronger hydrogen bond for the Schiff base than BR as judged from the frequency difference between their C[double bond]NH and C[double bond]ND stretches. We now find that replacement of the 10 amino acids of BR with ppR (BR/ppR) does not weaken the hydrogen bond of the Schiff base. Rather, the hydrogen bond in BR/ppR is stronger than that in the native ppR. We conclude that the principal factor of the smaller than expected opsin shift in BR/ppR is the strong association of the Schiff base with the surrounding counterion complex.
authors: Laboratory of Biophysical Chemistry, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan.
authors: Kazumi
authors: K
authors: Shimono
authors: Yuji
authors: Y
authors: Furutani
authors: Hideki
authors: H
authors: Kandori
authors: Naoki
authors: N
authors: Kamo
mesh_terms: D000577:Amides / Q000737:chemistry; D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics*; D001436:Bacteriorhodopsins / Q000737:chemistry*; D001665:Binding Sites / Q000235:genetics; D002338:Carotenoids / Q000737:chemistry* / Q000235:genetics*; D005615:Freezing; D025602:Halorhodopsins; D006860:Hydrogen Bonding; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D012172:Retinaldehyde / Q000737:chemistry*; D017299:Rod Opsins / Q000737:chemistry; D012545:Schiff Bases; D025603:Sensory Rhodopsins; D017550:Spectroscopy, Fourier Transform Infrared
chemical_list: D000577:Amides; D019843:Archaeal Proteins; D025602:Halorhodopsins; D017299:Rod Opsins; D012545:Schiff Bases; D025603:Sensory Rhodopsins; C061508:sensory rhodopsin II protein, archaeal; D002338:Carotenoids; D001436:Bacteriorhodopsins; D012172:Retinaldehyde