Preparation of a soluble 58 kDa-3-hydroxy-3-methylglutaryl CoA reductase from liver microsomes and its inhibition by ethoxysilatrane, a hypocholesterolemic compound.
On repeated thawing at room temperature of frozen preparations of heavy microsomes from rat livers, HMGCoA reductase activity was solubilized due to limited proteolysis. This soluble enzyme was partially purified by fractionation with ammonium sulfate and filtration on Sephacryl S-200 column. The active enzyme was coeluted with a major 92 kDa-protein and was identified as a 58 kDa-protein after separation by SDS-PAGE and immunoblotting. Ethoxysilatrane, a hypocholesterolemic compound, which decreased the liver-microsomal activity of HMGCoA reductase on intra-peritonial treatment of animals, showed little effect on the enzyme activity with isolated microsomes or the 50 kDa-soluble enzyme when added in the assay. But it was able to inhibit the activity of the soluble 58 kDa-enzyme in a concentration-dependent, reversible manner. Cholesterol and an oxycholesterol were without effect whereas chlorophenoxyisobutyrate and ubiquinone showed small inhibition under these conditions. The extra region that links the active site domain (50 kDa protein) to the membrane, present in the 58 kDa-protein appears to be involved in mediating the inhibition by silatrane.
Omkumar
R V
RV
Department of Biochemistry, Indian Institute of Science, Bangalore.
Mehta
P P
PP
Kurup
C K
CK
Ramasarma
T
T
D000818:Animals; D000924:Anticholesteremic Agents / Q000494:pharmacology*; D015153:Blotting, Western; D001643:Bridged Bicyclo Compounds / Q000494:pharmacology*; D019086:Bridged Bicyclo Compounds, Heterocyclic; D002850:Chromatography, Gel; D006903:Hydroxymethylglutaryl CoA Reductases / Q000302:isolation & purification / Q000378:metabolism*; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D017646:Organosilicon Compounds; D051381:Rats; D011919:Rats, Inbred Strains; D012825:Silicon / Q000494:pharmacology*; D012995:Solubility
D000924:Anticholesteremic Agents; D001643:Bridged Bicyclo Compounds; D019086:Bridged Bicyclo Compounds, Heterocyclic; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; D017646:Organosilicon Compounds; C015503:silatrane; C042292:1-ethoxysilatrane; D006903:Hydroxymethylglutaryl CoA Reductases; D012825:Silicon