Source:http://linkedlifedata.com/resource/pubmed/id/11418092
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rdf:type | |
lifeskim:mentions | |
pubmed:issue |
1
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pubmed:dateCreated |
2001-6-21
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pubmed:abstractText |
This investigation focused on the kinetics of cyanide binding to oxidized and reduced cytochrome d in Salmonella typhimurium intact cells, spheroplasts, membrane fragments and solubilized enzyme, and on the effect of pH on this binding. Cyanide bound to the oxidized form of cytochrome d under all experimental conditions, inducing a trough at 649 nm in the oxidized-cyanide-minus-oxidized difference absorption spectra. V(max) of cyanide binding to oxidized cytochrome d at pH 7.0 was 14.0+/-2.0 pmol/min/mg protein (prot.) in intact cells, 37.0+/-3.5 pmol/min/mg prot. in spheroplasts, 125.0+/-6.0 pmol/min/mg prot. in membrane fragments, and 538.0+/-8.5 pmol/min/mg prot. in solubilized cytochrome d. The pseudo-first order rate constants were 0.004 s(-1) for intact cells, 0.005 s(-1) for spheroplasts, 0.007 s(-1) for membrane fragments and 0.025 s(-1) for the solubilized enzyme. The V(max) value was highest at pH 7.0 for intact cells and solubilized cytochrome d and at pH 8.0 for both spheroplasts and membrane fragments. The K(s) of binding at pH 7.0 was around 4 mM in intact cells, spheroplasts and membrane fragments, but was 10.5 mM in solubilized cytochrome d. This difference between the K(s) values suggested a change in conformation, upon solubilization, leading to a decrease in the affinity of cyanide for the solubilized enzyme. The K(s) value was nearly the same at all pH investigated (pH 5-10). Cyanide was found to also bind to the reduced form of cytochrome d in membrane fragments (K(s)=18+/-3 mM, V(max)=377+/-28 pmol/min/mg prot. at pH 7) and the solubilized enzyme (K(s)=18+/-1.2 mM, V(max)=649+/-45 pmol/min/mg prot. at pH 7) with a lower affinity of cyanide for the reduced cytochrome d than for the oxidized enzyme. Pseudo-first order rate constants were 0.025 s(-1) and 0.042 s(-1) respectively for membrane fragments and solubilized enzyme. The value of V(max) for cyanide binding to the reduced cytochrome d, whether membrane-bound or solubilized, increased slightly with pH (for pH 6-10) while the K(s) value dropped significantly with increasing pH. The pH dependence observed here might be interpretable as a possible role for conformational transition associated with energy transduction. Finally, this investigation pointed to the influence of the microenvironment of a protein within the cell on its reactivity.
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pubmed:language |
eng
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pubmed:journal | |
pubmed:citationSubset |
IM
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pubmed:chemical | |
pubmed:status |
MEDLINE
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pubmed:month |
Jul
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pubmed:issn |
0006-3002
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pubmed:author | |
pubmed:issnType |
Print
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pubmed:day |
2
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pubmed:volume |
1506
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
1-11
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pubmed:dateRevised |
2006-11-15
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pubmed:meshHeading |
pubmed-meshheading:11418092-Cyanides,
pubmed-meshheading:11418092-Cytochrome d Group,
pubmed-meshheading:11418092-Hydrogen-Ion Concentration,
pubmed-meshheading:11418092-Kinetics,
pubmed-meshheading:11418092-Octoxynol,
pubmed-meshheading:11418092-Oxidation-Reduction,
pubmed-meshheading:11418092-Potassium Cyanide,
pubmed-meshheading:11418092-Salmonella typhimurium,
pubmed-meshheading:11418092-Spectrophotometry,
pubmed-meshheading:11418092-Spheroplasts,
pubmed-meshheading:11418092-Subcellular Fractions
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pubmed:year |
2001
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pubmed:articleTitle |
The binding of cyanide to cytochrome d in intact cells, spheroplasts, membrane fragments and solubilized enzyme from Salmonella typhimurium.
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pubmed:affiliation |
Institute of Biochemistry and Biophysics, University of Tehran, Tehran, Iran. keyhanie@ibb.ut.ac.ir
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pubmed:publicationType |
Journal Article,
Research Support, Non-U.S. Gov't
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