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PMID: 11425321 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

Chromophore structure in lumirhodopsin and metarhodopsin I by time-resolved resonance Raman microchip spectroscopy.

Biochemistry ·Vol. 40 ·No. 26 ·2001-07-03 ·Pages 7929-36

Pan D, Mathies RA

Abstract

Time-resolved resonance Raman microchip flow experiments have been performed on the lumirhodopsin (Lumi) and metarhodopsin I (Meta I) photointermediates of rhodopsin at room temperature to elucidate the structure of the chromophore in each species as well as changes in protein-chromophore interactions. Transient Raman spectra of Lumi and Meta I with delay times of 16 micros and 1 ms, respectively, are obtained by using a microprobe system to focus displaced pump and probe laser beams in a microfabricated flow channel and to detect the scattering. The fingerprint modes of both species are very similar and characteristic of an all-trans chromophore. Lumi exhibits a relatively normal hydrogen-out-of-plane (HOOP) doublet at 951/959 cm(-1), while Meta I has a single HOOP band at 957 cm(-1). These results suggest that the transitions from bathorhodopsin to Lumi and Meta I involve a relaxation of the chromophore to a more planar all-trans conformation and the elimination of the structural perturbation that uncouples the 11H and 12H wags in bathorhodopsin. Surprisingly, the protonated Schiff base C=N stretching mode in Lumi (1638 cm(-1)) is unusually low compared to those in rhodopsin and bathorhodopsin, and the C=ND stretching mode shifts down by only 7 cm(-1) in D2O buffer. This indicates that the Schiff base hydrogen bonding is dramatically weakened in the bathorhodopsin to Lumi transition. However, the C=N stretching mode in Meta I is found at 1654 cm(-1) and exhibits a normal deuteration-induced downshift of 24 cm(-1), identical to that of the all-trans protonated Schiff base. The structural relaxation of the chromophore-protein complex in the bathorhodopsin to Lumi transition thus appears to drive the Schiff base group out of its hydrogen-bonded environment near Glu113, and the hydrogen bonding recovers to a normal solvated PSB value but presumably a different hydrogen bond acceptor with the formation of Meta I.

MeSH Terms
Animals Cattle Hydrogen Bonding Photochemistry Protein Conformation Retinoids/chemistry Rhodopsin/analogs & derivatives,chemistry Schiff Bases/chemistry Spectrum Analysis, Raman/instrumentation,methods Temperature
Chemicals
Retinoids Schiff Bases lumirhodopsin retinylidene chromophore metarhodopsins Rhodopsin
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Pan D
Department of Chemistry, University of California, Berkeley, California 94720, USA.
Mathies R A
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Article Info
Journal
Biochemistry
Abbr.
Biochemistry
ISSN
0006-2960
DOI
10.1021/bi010670x
Published
2001-07-03
Pages
7929-36
Language
English
Region
United States
NLM ID
0370623
PMCID
PMC1404555
Subset
IM
Grants
NEI NIH HHS · R01 EY002051 · United States
NEI NIH HHS · R37 EY002051 · United States
NEI NIH HHS · EY02051 · United States
NEI NIH HHS · R01 EY002051-25 · United States
External Links
PubMed source DOI: 10.1021/bi010670x PMC Full Text View journal details
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