Mitochondrial citrate synthase is immobilized in vivo

Abstract

The enzymes of the tricarboxylic acid cycle in the mitochondrial matrix are proposed to form a multienzyme complex, in which there is channeling of substrates between enzyme active sites. However no direct evidence has been obtained in vivo for the involvement of these enzymes in such a complex. We have labeled the tricarboxylic acid cycle enzyme, citrate synthase 1, in the yeast Saccharomyces cerevisiae, by biosynthetic incorporation of 5-fluorotryptophan. Comparison of the 19F NMR resonance intensities from the labeled enzyme in the intact cell and in cell-free lysates indicated that the enzyme is motionally restricted in vivo, consistent with its participation in a multienzyme complex.