Femtosecond and picosecond dynamics of recombinant bacteriorhodopsin primary reactions compared to the native protein in trimeric and monomeric forms

O. A. Smitienko; O. V. Nekrasova; A. V. Kudriavtsev; M. A. Yakovleva; I. V. Shelaev; F. E. Gostev; D. A. Dolgikh; I. B. Kolchugina; V. A. Nadtochenko; M. P. Kirpichnikov; T. B. Feldman; M. A. Ostrovsky

doi:10.1134/S0006297917040113

Femtosecond and picosecond dynamics of recombinant bacteriorhodopsin primary reactions compared to the native protein in trimeric and monomeric forms

Authors: Smitienko O.A.¹, Nekrasova O.V.²^,3, Kudriavtsev A.V.¹^,3, Yakovleva M.A.¹, Shelaev I.V.⁴, Gostev F.E.⁴, Dolgikh D.A.²^,3^,5, Kolchugina I.B.³, Nadtochenko V.A.⁴, Kirpichnikov M.P.²^,3, Feldman T.B.¹^,3^,5, Ostrovsky M.A.¹^,3
Affiliations:
1. Emanuel Institute of Biochemical Physics
2. Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry
3. Lomonosov Moscow State University
4. Semenov Institute of Chemical Physics
5. Pirogov Russian National Research Medical University
Issue: Vol 82, No 4 (2017)
Pages: 490-500
Section: Article
URL: https://journals.rcsi.science/0006-2979/article/view/151351
DOI: https://doi.org/10.1134/S0006297917040113
ID: 151351

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Abstract

Photochemical reaction dynamics of the primary events in recombinant bacteriorhodopsin (bR_rec) was studied by femtosecond laser absorption spectroscopy with 25-fs time resolution. bR_rec was produced in an Escherichia coli expression system. Since bR_rec was prepared in a DMPC–CHAPS micelle system in the monomeric form, its comparison with trimeric and monomeric forms of the native bacteriorhodopsin (bR_trim and bR_mon, respectively) was carried out. We found that bR_rec intermediate I (excited state of bR) was formed in the range of 100 fs, as in the case of bR_trim and bR_mon. Further processes, namely the decay of the excited state I and the formation of intermediates J and K of bR_rec, occurred more slowly compared to bR_trim, but similarly to bR_mon. The lifetime of intermediate I, judging from the signal of ΔA_ESA(470-480 nm), was 0.68 ps (78%) and 4.4 ps (22%) for bR_rec, 0.52 ps (73%) and 1.7 ps (27%) for bR_mon, and 0.45 ps (90%) and 1.75 ps (10%) for bR_trim. The formation time of intermediate K, judging from the signal of ΔA_GSA(625-635 nm), was 13.5 ps for bR_rec, 9.8 ps for bR_mon, and 4.3 ps for bR_trim. In addition, there was a decrease in the photoreaction efficiency of bR_rec and bR_mon as seen by a decrease in absorbance in the differential spectrum of the intermediate K by ~14%. Since photochemical properties of bR_rec are similar to those of the monomeric form of the native protein, bR_rec and its mutants can be considered as a basis for further studies of the mechanism of bacteriorhodopsin functioning.

Keywords

bacteriorhodopsin, recombinant protein, primary reactions, femtosecond absorption laser spectroscopy

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Femtosecond and picosecond dynamics of recombinant bacteriorhodopsin primary reactions compared to the native protein in trimeric and monomeric forms

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Abstract

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About the authors

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