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PMID: 10692336 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Unraveling photoexcited conformational changes of bacteriorhodopsin by time resolved electron paramagnetic resonance spectroscopy.

Biophysical journal ·Vol. 78 ·No. 3 ·2000-03-00 ·Pages 1519-30

Rink T, Pfeiffer M, Oesterhelt D, Gerwert K, Steinhoff HJ

Abstract

By means of time-resolved electron paramagnetic resonance (EPR) spectroscopy, the photoexcited structural changes of site-directed spin-labeled bacteriorhodopsin are studied. A complete set of cysteine mutants of the C-D loop, positions 100-107, and of the E-F loop, including the first alpha-helical turns of helices E and F, positions 154-171, was modified with a methanethiosulfonate spin label. The EPR spectral changes occurring during the photocycle are consistent with a small movement of helix C and an outward tilt of helix F. These helix movements are accompanied by a rearrangement of the E-F loop and of the C-terminal turn of helix E. The kinetic analysis of the transient EPR data and the absorbance changes in the visible spectrum reveals that the conformational change occurs during the lifetime of the M intermediate. Prominent rearrangements of nitroxide side chains in the vicinity of D96 may indicate the preparation of the reprotonation of the Schiff base. All structural changes reverse with the recovery of the bacteriorhodopsin initial state.

MeSH Terms
Amino Acid Sequence Amino Acid Substitution Bacteriorhodopsins/chemistry,radiation effects Electron Spin Resonance Spectroscopy/methods Models, Molecular Mutagenesis, Site-Directed Photochemistry Protein Conformation/radiation effects Protein Structure, Secondary Recombinant Proteins/chemistry,radiation effects Spin Labels Time Factors
Chemicals
Recombinant Proteins Spin Labels Bacteriorhodopsins
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Rink T
Lehrstuhl für Biophysik, Ruhr-Universität Bochum, 44801 Bochum, Germany.
Pfeiffer M
Oesterhelt D
Gerwert K
Steinhoff H J
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Article Info
Journal
Biophysical journal
Abbr.
Biophys J
ISSN
0006-3495
DOI
10.1016/S0006-3495(00)76704-X
Published
2000-03-00
Pages
1519-30
Language
English
Region
United States
NLM ID
0370626
PMCID
PMC1300749
Subset
IM
External Links
PubMed source DOI: 10.1016/S0006-3495(00)76704-X PMC Full Text View journal details
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