title: Femtosecond and Picosecond Dynamics of Recombinant Bacteriorhodopsin Primary Reactions Compared to the Native Protein in Trimeric and Monomeric Forms.
abstract: Photochemical reaction dynamics of the primary events in recombinant bacteriorhodopsin (bRrec) was studied by femtosecond laser absorption spectroscopy with 25-fs time resolution. bRrecwas produced in an Escherichia coli expression system. Since bRrecwas prepared in a DMPC-CHAPS micelle system in the monomeric form, its comparison with trimeric and monomeric forms of the native bacteriorhodopsin (bRtrimand bRmon, respectively) was carried out. We found that bRrecintermediate I (excited state of bR) was formed in the range of 100 fs, as in the case of bRtrimand bRmon. Further processes, namely the decay of the excited state I and the formation of intermediates J and K of bRrec, occurred more slowly compared to bRtrim, but similarly to bRmon. The lifetime of intermediate I, judging from the signal of ΔAESA(470-480 nm), was 0.68 ps (78%) and 4.4 ps (22%) for bRrec, 0.52 ps (73%) and 1.7 ps (27%) for bRmon, and 0.45 ps (90%) and 1.75 ps (10%) for bRtrim. The formation time of intermediate K, judging from the signal of ΔAGSA(625-635 nm), was 13.5 ps for bRrec, 9.8 ps for bRmon, and 4.3 ps for bRtrim. In addition, there was a decrease in the photoreaction efficiency of bRrecand bRmonas seen by a decrease in absorbance in the differential spectrum of the intermediate K by ~14%. Since photochemical properties of bRrecare similar to those of the monomeric form of the native protein, bRrecand its mutants can be considered as a basis for further studies of the mechanism of bacteriorhodopsin functioning.
authors.affiliation: Emanuel Institute of Biochemical Physics, Russian Academy of Sciences, Moscow, 119334, Russia. djolia@gmail.com.
authors.forename: O A
authors.initials: OA
authors.lastname: Smitienko
authors.forename: O V
authors.initials: OV
authors.lastname: Nekrasova
authors.forename: A V
authors.initials: AV
authors.lastname: Kudriavtsev
authors.forename: M A
authors.initials: MA
authors.lastname: Yakovleva
authors.forename: I V
authors.initials: IV
authors.lastname: Shelaev
authors.forename: F E
authors.initials: FE
authors.lastname: Gostev
authors.forename: D A
authors.initials: DA
authors.lastname: Dolgikh
authors.forename: I B
authors.initials: IB
authors.lastname: Kolchugina
authors.forename: V A
authors.initials: VA
authors.lastname: Nadtochenko
authors.forename: M P
authors.initials: MP
authors.lastname: Kirpichnikov
authors.forename: T B
authors.initials: TB
authors.lastname: Feldman
authors.forename: M A
authors.initials: MA
authors.lastname: Ostrovsky
mesh_terms: D001436:Bacteriorhodopsins / Q000737:chemistry*; D001704:Biopolymers / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D011994:Recombinant Proteins / Q000737:chemistry; D013056:Spectrophotometry, Ultraviolet
chemical_list: D001704:Biopolymers; D011994:Recombinant Proteins; D001436:Bacteriorhodopsins