| The role of back-reactions and proton uptake during the N----O transition in bacteriorhodopsin's photocycle: a kinetic resonance Raman study. | |
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MedLine Citation:
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PMID: 2169875 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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The kinetics of bacteriorhodopsin's photocycle have been analyzed at pH 5, 6, 7, 8, and 8.6 by using time-resolved resonance Raman spectroscopy. The concentrations of the various intermediates as a function of time were determined by following their resonance Raman intensities using 502-nm (L550, N550, BR568), 458-nm (M412), and 752-nm (O640) excitation. The spectral contributions to the pump + probe data from each intermediate were quantitatively separated by least-squares decomposition. These relative concentrations were then converted to absolute concentrations by using a conservation of molecules constraint. This enabled the unambiguous refinement of a variety of kinetic models to find the simplest one that accurately describes the data. The kinetic data, including the biphasic decay of L550 and M412, are best reproduced by a sequential scheme including back-reactions (BR----L----M----N----O----BR). In addition, the kinetics of the L----M and N----O steps are found to be pH-dependent. Both the forward and reverse rate constants connecting L550 and M412 increase with pH, confirming earlier proposals of catalyzed Schiff base deprotonation at alkaline pH. Below pH 7, the N550----O640 rate constant is independent of pH, but it decreases linearly with pH above 7. This indicates that the protein must pick up a proton during the N550----O640 transition and that this process becomes rate determining above pH 7. There must, therefore, be an intermediate between N550 and O640 which we denote as N+550. A molecular graphics model is presented which incorporates these observations into a mechanism for proton pumping. |
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Authors:
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J B Ames; R A Mathies |
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Publication Detail:
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Type: Comparative Study; Journal Article; Research Support, U.S. Gov't, Non-P.H.S.; Research Support, U.S. Gov't, P.H.S. |
Journal Detail:
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Title: Biochemistry Volume: 29 ISSN: 0006-2960 ISO Abbreviation: Biochemistry Publication Date: 1990 Aug |
Date Detail:
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Created Date: 1990-11-20 Completed Date: 1990-11-20 Revised Date: 2007-11-14 |
Medline Journal Info:
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Nlm Unique ID: 0370623 Medline TA: Biochemistry Country: UNITED STATES |
Other Details:
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Languages: eng Pagination: 7181-90 Citation Subset: IM |
Affiliation:
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Department of Chemistry, University of California, Berkeley 94720. |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Bacteriorhodopsins
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chemistry,
metabolism*,
radiation effects Halobacterium / analysis Hydrogen-Ion Concentration Kinetics Photochemistry Protein Conformation Protons Spectrum Analysis, Raman |
| Grant Support | |
ID/Acronym/Agency:
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GM 27057/GM/NIGMS NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/Protons; 53026-44-1/Bacteriorhodopsins |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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