title: Enzyme inhibition during the conversion of squalene to cholesterol.
abstract: Two separate enzymatic assays were developed in order to test the selectivity of inhibitors in cholesterol biosynthesis. One assay detects inhibition of delta 5.7-sterol delta 7-reductase, the enzyme involved in the conversion of 7-dehydrocholesterol to cholesterol. Delta 5.7-Sterol delta 7-reductase was inhibited by both RPR 101821, a protonated cyclohexylamine, and BM 15.766, a piperazine derivative, with IC50 values of 1 microM. The second assay detects accumulation of any of five intermediates (squalene oxide, squalene dioxide, lanosterol, desmosterol, and 7-dehydrocholesterol) upon inhibition of enzymes catalyzing reactions in the conversion of squalene to cholesterol. In this assay, inhibition data were most accurate when control assays exhibited a conversion of squalene to cholesterol in the order of 50%. The time required to attain 50% conversion of squalene to cholesterol was 6 h. Given a high inhibitor to substrate concentration ratio and the possible values of Ki, kon, and koff for the reaction between enzymes and inhibitor to form enzyme-inhibitor complexes, it was predicted that in the presence of inhibitors, intermediate accumulation could still be observed after 6 h incubation. The experimental results were in agreement with this prediction.
mesh_terms: D000818:Animals; D001583:Benzoxazoles / Q000494:pharmacology; D002784:Cholesterol / Q000378:metabolism*; D003514:Cyclohexylamines / Q000494:pharmacology; D003684:Dehydrocholesterols / Q000302:isolation & purification / Q000378:metabolism; D003897:Desmosterol / Q000302:isolation & purification / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D013186:Farnesyl-Diphosphate Farnesyltransferase / Q000037:antagonists & inhibitors*; D007810:Lanosterol / Q000302:isolation & purification / Q000378:metabolism; D008297:Male; D010088:Oxidoreductases / Q000037:antagonists & inhibitors; D044925:Oxidoreductases Acting on CH-CH Group Donors; D010879:Piperazines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D013185:Squalene / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D001583:Benzoxazoles; D003514:Cyclohexylamines; D003684:Dehydrocholesterols; D004791:Enzyme Inhibitors; D010879:Piperazines; C096297:RPR 101821; C033144:lathosterol delta-5-dehydrogenase; D007810:Lanosterol; D003897:Desmosterol; D013185:Squalene; C047457:BM 15766; D002784:Cholesterol; C016705:7-dehydrocholesterol; D010088:Oxidoreductases; D044925:Oxidoreductases Acting on CH-CH Group Donors; D013186:Farnesyl-Diphosphate Farnesyltransferase