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Pathways of proton release in the bacteriorhodopsin photocycleThe pH dependencies of the rate constants in the photocycles of recombinant D96N and D115N/D96N bacteriorhodopsins were determined from time-resolved difference spectra between 70 ns and 420 ms after photoexcitation. The results were consistent with the model suggested earlier for proteins containing D96N substitution: BR hv----K----L----M1----M2----BR. Only the M2----M1 back-reaction was pH-dependent: its rate increased with increasing [H+] between pH 5 and 8. We conclude from quantitative analysis of this pH dependency that its reverse, the M1----M2 reaction, is linked to the release of a proton from a group with a pKa = 5.8. This suggests a model for wild-type bacteriorhodopsin in which at pH greater than 5.8 the transported proton is released on the extracellular side from this as yet unknown group and on the 100-microseconds time scale, but at pH less than 5.8, the proton release occurs from another residue and later in the photocycle most likely directly from D85 during the O----BR reaction. We postulate, on the other hand, that proton uptake on the cytoplasmic side will be by D96 and during the N----O reaction regardless of pH. The proton kinetics as measured with indicator dyes confirmed the unique prediction of this model: at pH greater than 6, proton release preceded proton uptake, but at pH less than 6, the release was delayed until after the uptake. The results indicated further that the overall M1----M2 reaction includes a second kinetic step in addition to proton release; this is probably the earlier postulated extracellular-to-cytoplasmic reorientation switch in the proton pump.
Document ID
20050000607
Acquisition Source
Legacy CDMS
Document Type
Reprint (Version printed in journal)
External Source(s)
Authors
Zimanyi, L.
(University of California Irvine 92717)
Varo, G.
Chang, M.
Ni, B.
Needleman, R.
Lanyi, J. K.
Date Acquired
August 22, 2013
Publication Date
September 15, 1992
Publication Information
Publication: Biochemistry
Volume: 31
Issue: 36
ISSN: 0006-2960
Subject Category
Life Sciences (General)
Funding Number(s)
CONTRACT_GRANT: GM 29498
Distribution Limits
Public
Copyright
Other
Keywords
NASA Discipline Exobiology
Non-NASA Center

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