Document Detail


Two groups control light-induced Schiff base deprotonation and the proton affinity of Asp85 in the Arg82 his mutant of bacteriorhodopsin.
MedLine Citation:
PMID:  10545374     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Arg(82) is one of the four buried charged residues in the retinal binding pocket of bacteriorhodopsin (bR). Previous studies show that Arg(82) controls the pK(a)s of Asp(85) and the proton release group and is essential for fast light-induced proton release. To further investigate the role of Arg(82) in light-induced proton pumping, we replaced Arg(82) with histidine and studied the resulting pigment and its photochemical properties. The main pK(a) of the purple-to-blue transition (pK(a) of Asp(85)) is unusually low in R82H: 1.0 versus 2.6 in wild type (WT). At pH 3, the pigment is purple and shows light and dark adaptation, but almost no light-induced Schiff base deprotonation (formation of the M intermediate) is observed. As the pH is increased from 3 to 7 the M yield increases with pK(a) 4.5 to a value approximately 40% of that in the WT. A transition with a similar pK(a) is observed in the pH dependence of the rate constant of dark adaptation, k(da). These data can be explained, assuming that some group deprotonates with pK(a) 4.5, causing an increase in the pK(a) of Asp(85) and thus affecting k(da) and the yield of M. As the pH is increased from 7 to 10.5 there is a further 2.5-fold increase in the yield of M and a decrease in its rise time from 200 micros to 75 micros with pK(a) 9. 4. The chromophore absorption band undergoes a 4-nm red shift with a similar pK(a). We assume that at high pH, the proton release group deprotonates in the unphotolyzed pigment, causing a transformation of the pigment into a red-shifted "alkaline" form which has a faster rate of light-induced Schiff base deprotonation. The pH dependence of proton release shows that coupling between Asp(85) and the proton release group is weakened in R82H. The pK(a) of the proton release group in M is 7.2 (versus 5.8 in the WT). At pH < 7, most of the proton release occurs during O --> bR transition with tau approximately 45 ms. This transition is slowed in R82H, indicating that Arg(82) is important for the proton transfer from Asp(85) to the proton release group. A model describing the interaction of Asp(85) with two ionizable residues is proposed to describe the pH dependence of light-induced Schiff base deprotonation and proton release.
Authors:
E S Imasheva; S P Balashov; T G Ebrey; N Chen; R K Crouch; D R Menick
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural; Research Support, U.S. Gov't, Non-P.H.S.    
Journal Detail:
Title:  Biophysical journal     Volume:  77     ISSN:  1542-0086     ISO Abbreviation:  Biophys. J.     Publication Date:  1999 Nov 
Date Detail:
Created Date:  2010-08-30     Completed Date:  2010-12-03     Revised Date:  2012-05-07    
Medline Journal Info:
Nlm Unique ID:  0370626     Medline TA:  Biophys J     Country:  United States    
Other Details:
Languages:  eng     Pagination:  2750-63     Citation Subset:  IM    
Affiliation:
Center for Biophysics and Computational Biology and Department of Cell and Structural Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
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MeSH Terms
Descriptor/Qualifier:
Absorption
Adaptation, Physiological
Amino Acid Substitution
Bacteriorhodopsins / chemistry*,  genetics,  metabolism*
Biological Transport
Color
Darkness
Extracellular Space / metabolism
Halobacterium salinarum / cytology,  metabolism,  physiology,  radiation effects
Hydrogen-Ion Concentration
Kinetics
Light*
Models, Molecular
Mutant Proteins / chemistry*,  genetics,  metabolism*
Pigments, Biological / chemistry,  metabolism
Protein Conformation
Protons*
Schiff Bases / metabolism*
Stereoisomerism
Grant Support
ID/Acronym/Agency:
GM52023/GM/NIGMS NIH HHS
Chemical
Reg. No./Substance:
0/Mutant Proteins; 0/Pigments, Biological; 0/Protons; 0/Schiff Bases; 53026-44-1/Bacteriorhodopsins
Comments/Corrections

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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