Studies on the citryl-CoA-dependent inhibition of citrate-synthase with source variants from baker's yeast, Escherichia coli and Sulfolobus solfataricus.
1) Citrate synthase from pig heart has previously been shown to display complex kinetic characteristics in the reactions with citryl-CoA, resulting in inhibition. The synthase from another eukaryotic source, baker's yeast, yields the same complex kinetics. 2) Synthases from a Gram-negative prokaryote, E. coli, and from an archaebacterium, S. solfataricus, catalyse the reactions of citryl-CoA in kinetics of the Michaelis-Menten type. A comparison of the rates of citryl-CoA hydrolysis (V') and physiological reaction (V), determined with these enzymes, corresponds to ratios of V'/V approximately 1 and approximately 2, respectively. Thus, and for the first time, there is no reason left to doubt the intermediate formation of citryl-CoA in the physiological reaction. 3) The complex kinetics indicated under 1) are related to efficient formation of citrate from citryl-CoA-derived acetyl-CoA and oxaloacetate in the presence of NADH and malate dehydrogenase. These conditions are not met by the enzymes from E. coli, S. solfataricus and by proteolytically nicked synthase species from pig heart. All these enzyme variants have low affinities to either one or both of the physiological substrates. Consistent with earlier ideas, the results indicate that the inhibition mechanism is related to high affinities of the enzyme for both acetyl-CoA and oxaloacetate.
Löhlein-Werhahn
G
G
Institut für Physiologische Chemie der Technischen Universität München.
Goepfert
P
P
Kollmann-Koch
A
A
Eggerer
H
H
D000214:Acyl Coenzyme A / Q000494:pharmacology*; D001105:Archaea / Q000201:enzymology*; D001419:Bacteria / Q000201:enzymology*; D002950:Citrate (si)-Synthase / Q000037:antagonists & inhibitors*; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D010071:Oxaloacetates / Q000494:pharmacology; D007652:Oxo-Acid-Lyases / Q000037:antagonists & inhibitors*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013045:Species Specificity
D000214:Acyl Coenzyme A; D010071:Oxaloacetates; C029890:citryl-coenzyme A; D002950:Citrate (si)-Synthase; D007652:Oxo-Acid-Lyases