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Chromophore structure in bacteriorhodopsin's N intermediate: implications for the proton-pumping mechanism

Journal Article · · Biochemistry; (United States)
DOI:https://doi.org/10.1021/bi00418a064· OSTI ID:6786610
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By elevating the pH to 9.5 in 3 M KCl, the concentration of the N intermediate in the bacteriorhodopsin photocycle has been enhanced, and time-resolved resonance Raman spectra of this intermediate have been obtained. Kinetic Raman measurements show that N appears with a half-time of 4 +/- 2 ms, which agrees satisfactorily with our measured decay time of the M412 intermediate (2 +/- 1 ms). This argues that M412 decays directly to N in the light-adapted photocycle. The configuration of the chromophore about the C13 = C14 bond was examined by regenerating the protein with (12,14-2H)retinal. The coupled C12-2H + C14-2H rock at 946 cm-1 demonstrates that the chromophore in N is 13-cis. The shift of the 1642-cm-1 Schiff base stretching mode to 1618 cm-1 in D2O indicates that the Schiff base linkage to the protein is protonated. The insensitivity of the 1168-cm-1 C14-C15 stretching mode to N-deuteriation establishes a C = N anti (trans) Schiff base configuration. The high frequency of the C14-C15 stretching mode as well as the frequency of the 966-cm-1 C14-2H-C15-2H rocking mode shows that the chromophore is 14-s-trans. Thus, N contains a 13-cis, 14-s-trans, 15-anti protonated retinal Schiff base.(ABSTRACT TRUNCATED AT 250 WORDS)

Research Organization:
Univ. of California, Berkeley (USA)
OSTI ID:
6786610
Journal Information:
Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 27:18; ISSN BICHA
Country of Publication:
United States
Language:
English

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Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
BARYONS
CARBOXYLIC ACID ESTERS
CHEMISTRY
DEUTERIUM
ELEMENTARY PARTICLES
ESTERS
FERMIONS
HADRONS
HYDROGEN ISOTOPES
IMINES
ISOTOPE APPLICATIONS
ISOTOPES
LIGHT NUCLEI
MOLECULAR STRUCTURE
NUCLEI
NUCLEONS
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PHOTOCHEMISTRY
PIGMENTS
PROTEINS
PROTONS
RAMAN SPECTRA
RETINOIC ACID
RHODOPSIN
SCHIFF BASES
SPECTRA
STABLE ISOTOPES
TRACER TECHNIQUES