Phosphorylation of 3-hydroxy-3-methylglutaryl coenzyme A reductase by microsomal 3-hydroxy-3-methylglutaryl coenzyme A reductase kinase.
Microsomal 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase kinase has been purified to apparent homogeneity by a process involving the following steps: solubilization from microsomes and chromatography on Affi-Gel Blue, phosphocellulose, Bio-Gel A 1.5m, and agarose-hexane-ATP. The apparent Mr of the purified enzyme as judged by gel-filtration chromatography is 205,000 and by sodium dodecyl sulfate-gel electrophoresis is 105,000. Immunoprecipitation of homogeneous reductase phosphorylated by reductase kinase and [gamma-32P]ATP produces a unique band containing 32P bound to protein which migrates at the same Rf as the reductase subunit. Incubation of 32P-labeled HMG-CoA reductase with reductase phosphatase results in a time-dependent loss of protein-bound 32P radioactivity, as well as an increase in enzymic activity. Reductase kinase, when incubated with ATP, undergoes autophosphorylation, and a simultaneous increase in its enzymatic activity is observed. Tryptic treatment of immunoprecipitated, 32P-labeled HMG-CoA reductase phosphorylated with reductase kinase produces only one 32P-labeled phosphopeptide with the same Rf as one of the two tryptic phosphopeptides that have been reported in a previous paper. The possible existence of a second microsomal reductase kinase is discussed.
D055372:AMP-Activated Protein Kinases; D000818:Animals; D001665:Binding Sites; D002845:Chromatography / Q000379:methods; D006903:Hydroxymethylglutaryl CoA Reductases / Q000378:metabolism*; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D009097:Multienzyme Complexes / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D051381:Rats; D011919:Rats, Inbred Strains; D012995:Solubility
D009097:Multienzyme Complexes; D006903:Hydroxymethylglutaryl CoA Reductases; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D055372:AMP-Activated Protein Kinases