Mevalonate utilization in Pseudomonas sp. M. Purification and characterization of an inducible 3-hydroxy-3-methylglutaryl coenzyme A reductase.
Pseudomonas sp. M grown on mevalonate as the sole source of carbon has 200- to 800-fold induced levels of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase. The enzyme, which was purified to a homogeneous state in 54% yield (final specific activity, 60.5 mumol of NAD+ reduced per min per mg of protein), converted R-mevalonate (Km = 0.15 mM) to S-HMG-CoA. Activity was sensitive to sulfhydryl modifying reagents. The apparent molecular weight of the holoenzyme was 178,000 and that of the subunit 43,000. The enzyme thus appears to be a tetramer. Comparison of a 23-residue amino-terminal sequence with the cDNA-derived sequence of Chinese hamster ovary cell HMG-CoA reductase showed little homology and antibody raised against the Pseudomonas enzyme did not appear to cross-react with rat liver HMG-CoA reductase. Addition of mevalonate to cells growing on glucose was followed by a rapid and biphasic induction of HMG-CoA reductase activity. During phase I, mevalonate or its catabolites may accumulate in intact cells of Pseudomonas sp. M and acetoacetate, a competitive inhibitor of HMG-CoA reductase (Ki = 3.2 mM), may feedback inhibit the enzyme under these conditions.
D000596:Amino Acids / Q000032:analysis; D003470:Culture Media; D004790:Enzyme Induction; D006903:Hydroxymethylglutaryl CoA Reductases / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008798:Mevalonic Acid / Q000378:metabolism* / Q000494:pharmacology; D008970:Molecular Weight; D011549:Pseudomonas / Q000378:metabolism*; D013439:Sulfhydryl Reagents / Q000494:pharmacology
D000596:Amino Acids; D003470:Culture Media; D013439:Sulfhydryl Reagents; D006903:Hydroxymethylglutaryl CoA Reductases; D008798:Mevalonic Acid