Purification and Some Properties of Citrate Synthase from Penicillium spiculisporum

Citrate synthase has been purified 300 to 400‐fold to a maximum specific activity of 50—60 U/mg protein. It has similar properties to the majority of the eucaryotic citrate synthases which have been studied, although it is inhibited less by ATP. The K m for acetyl‐CoA is 30 μM and for oxaloacetate 10 μM. The apparent K m for acetyl‐CoA in the presence of 2.5 mM ATP is 50 μM and, in the presence of 5 mM ATP, 80 μM. The K i for ATP is 3—4 mM. The ATP inhibition is reduced by Mg 2+ . At 20 μM acetyl‐CoA 12% inhibition results from the transition from 5 mM AMP to 5 mM ATP in presence of a slight excess of Mg 2+ . The size of citrate synthase as estimated from gel‐filtration is about that of serum albumin. No close relationship between citrate synthase and 2‐decylcitrate synthase from Penicillium spiculisporum is apparent. If citrate is considered to be the ultimate precursor of decylcitrate, the extracted amount of citrate synthase is hardly sufficient to account for the observed production unless the enzyme operates at or near conditions of maximal velocity, which, in presence of adenine nucleotides, requires considerable effective substrate concentrations.