The synthesis of 3-nonaprenyl-4-hydroxybenzoate in rat liver mitochondria: The effect of Ca 2+, Mg 2+, chelators, and bacitracin on the activity of p-hydroxybenzoate-polyprenyl transferase

Abstract

The formation of the first intermediate in ubiquinone-9 biosynthesis, 3-nonaprenyl-4-hydroxybenzoate (NPHB), by the enzyme p-hydroxybenzoate:polyprenyl transferase, has been studied in isolated rat liver mitochondria using solanesol pyrophosphate and p-hydroxybenzoate as the substrates. Phosphate buffer (100 m m) is inhibitory but at 20 m m inhibition is not apparent compared to other buffers at the same concentration. With various buffers at low concentration (20 m m) both EDTA and Mg 2+ stimulate formation of NPHB while Ca 2+ inhibits. Release of Ca 2+ inhibition can be achieved by the addition of Mg 2+, or EDTA, or EGTA, with EGTA being less effective than EDTA. When Mg 2+, Ca 2+, and EDTA are present together, a two- to threefold increase in activity of the enzyme is observed. The antibiotic bacitracin inhibits the synthesis of NPHB and the inhibition is increased when divalent cations are present. EGTA is more effective than EDTA in overcoming inhibition due to bacitracin. The possibility that these effects are partially due to alteration of mitochondrial membrane conformation as well as a direct effect on the enzyme is evaluated. The possible role of polyprenylphosphates in mitochondrial membrane function is discussed.