Interactions of a photosensitive analog of cholesterol with hydroxymethyglutaryl-CoA reductase (NADPH) and acyl-CoA:cholesterol acyltransferase.
25-Azido[25-(3)H]norcholesten-3beta-ol, a radioactive photosensitive analog of cholesterol, was synthesized as a probe to study lipid-protein interactions [Stoffel, W. & Klotzbücher, R. (1978) Hoppe-Seyler's Z. Physiol Chem. 359, 199-209]. Upon UV irradiation a reactive nitrene is generated which can insert into carbon-hydrogen bonds of proteins to form covalent linkages. The compound has properties similar to those of cholesterol. When administered intravenously to rats it was transported to the liver, and within 4 hr it was found in all of the subcellular fractions, accounting for 8% of the microsomal sterol content. Concomitantly, it altered the activities of two membrane-bound enzymes. 3-Hydroxy-3-methylglutaryl (HMG)-CoA (NADPH) reductase [mevalonate:NADP(+) oxidoreductase (CoA-acylating), EC 1.1.1.34] activity was reduced to 20% of control values, and acyl-CoA:cholesterol acyltransferase (ACAT; acyl-CoA:cholesterol O-acyltransferase, EC 2.3.1.26) was stimulated at least 2-fold. Esterified products of the analog were found in both blood and liver, and no evidence was obtained for production of any hydroxylated or polar derivatives. UV irradiation of microsomes increased the radioactivity bound to the protein fraction and decreased it in the total lipid extract; in parallel a sharp decline in ACAT activity but unaltered HMG-CoA reductase activity was observed. These results are interpreted as a possible consequence of crosslinking of the sterol to the enzyme proteins and discussed in view of the evidence that HMG-CoA reductase is an extrinsic and ACAT an intrinsic membrane protein.
D000217:Acyltransferases / Q000378:metabolism*; D000818:Animals; D002784:Cholesterol / Q000031:analogs & derivatives* / Q000378:metabolism / Q000528:radiation effects; D006903:Hydroxymethylglutaryl CoA Reductases / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism; D007700:Kinetics; D008563:Membrane Lipids / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D008862:Microsomes, Liver / Q000378:metabolism / Q000528:radiation effects; D011485:Protein Binding; D051381:Rats; D002785:Sterol O-Acyltransferase / Q000378:metabolism*; D014466:Ultraviolet Rays
D008563:Membrane Lipids; D008565:Membrane Proteins; D002784:Cholesterol; D006903:Hydroxymethylglutaryl CoA Reductases; D000217:Acyltransferases; D002785:Sterol O-Acyltransferase