PMID:8955296
| Citation |
Kwon, O, Bhattacharyya, DK and Meganathan, R (1996) Menaquinone (vitamin K2) biosynthesis: overexpression, purification, and properties of o-succinylbenzoyl-coenzyme A synthetase from Escherichia coli. J. Bacteriol. 178:6778-81 |
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| Abstract |
The coenzyme A (CoA)- and ATP-dependent conversion of o-succinylbenzoic acid [OSB; 4-(2'-carboxyphenyl)-4-oxobutyric acid], to o-succinylbenzoyl-CoA is carried out by the enzyme o-succinylbenzoyl-CoA synthetase. o-Succinylbenzoyl-CoA is a key intermediate in the biosynthesis of menaquinone (vitamin K2) in both gram-negative and gram-positive bacteria. The enzyme has been overexpressed and purified to homogeneity. The purified enzyme was found to have a native molecular mass of 185 kDa as determined by gel filtration column chromatography on Sephacryl S-200. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis established a subunit molecular mass of 49 kDa. Thus, the enzyme is a homotetramer. The enzyme showed a pH optimum of 7.5 to 8.0 and a temperature optimum of 30 to 40 degrees C. The Km values for OSB, ATP, and CoA were 16, 73.5, and 360 microM, respectively. Of the various metal ions tested, Mg2+ was found to be the most effective in stimulating the enzyme activity. Studies with substrate analogs showed that neither benzoic acid nor benzoylpropionic acid (succinylbenzene) is a substrate for the enzyme. Thus, it appears that both the benzoyl carboxyl group and the succinyl side chain are required for activation of the aliphatic carboxyl group. |
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| Keywords |
Benzoates/metabolism; Cations/pharmacology; Escherichia coli/enzymology; Hydrogen-Ion Concentration; Kinetics; Magnesium/pharmacology; Molecular Weight; Spectrophotometry, Ultraviolet; Substrate Specificity; Succinate-CoA Ligases/chemistry; Succinate-CoA Ligases/genetics; Succinate-CoA Ligases/isolation & purification; Succinate-CoA Ligases/metabolism; Temperature; Transformation, Bacterial; Vitamin K/biosynthesis |
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