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Deriving the intermediate spectra and photocycle kinetics from time-resolved difference spectra of bacteriorhodopsin. The simpler case of the recombinant D96N proteinThe bacteriorhodopsin photocycle contains more than five spectrally distinct intermediates, and the complexity of their interconversions has precluded a rigorous solution of the kinetics. A representation of the photocycle of mutated D96N bacteriorhodopsin near neutral pH was given earlier (Varo, G., and J. K. Lanyi. 1991. Biochemistry. 30:5008-5015) as BRhv-->K<==>L<==>M1-->M2--> BR. Here we have reduced a set of time-resolved difference spectra for this simpler system to three base spectra, each assumed to consist of an unknown mixture of the pure K, L, and M difference spectra represented by a 3 x 3 matrix of concentration values between 0 and 1. After generating all allowed sets of spectra for K, L, and M (i.e., M1 + M2) at a 1:50 resolution of the matrix elements, invalid solutions were eliminated progressively in a search based on what is expected, empirically and from the theory of polyene excited states, for rhodopsin spectra. Significantly, the average matrix values changed little after the first and simplest of the search criteria that disallowed negative absorptions and more than one maximum for the M intermediate. We conclude from the statistics that during the search the solutions strongly converged into a narrow region of the multidimensional space of the concentration matrix. The data at three temperatures between 5 and 25 degrees C yielded a single set of spectra for K, L, and M; their fits are consistent with the earlier derived photocycle model for the D96N protein.
Document ID
20050000530
Acquisition Source
Legacy CDMS
Document Type
Reprint (Version printed in journal)
Authors
Zimanyi, L.
(University of California Irvine 92717)
Lanyi, J. K.
Date Acquired
August 22, 2013
Publication Date
January 1, 1993
Publication Information
Publication: Biophysical journal
Volume: 64
Issue: 1
ISSN: 0006-3495
Subject Category
Life Sciences (General)
Funding Number(s)
CONTRACT_GRANT: GM-29498
Distribution Limits
Public
Copyright
Other
Keywords
NASA Program Exobiology
NASA Discipline Exobiology
NASA Discipline Number 52-30
Non-NASA Center

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