| Summary: | For investigation of the reactions of cholesterol biosynthesis, a number of workers use the 10,000 g supernatant fraction (or similar preparations) obtained from cell-free homogenates of rat liver. We have found that esters of methyl sterol biosynthetic intermediates are formed by this crude source of enzymes. Esters of (C30-, C29-, C28-, and C27-sterol intermediates have been isolated by silicic acid chromatography of an acetone extract of incubation mixtures. Competition between ester formation and demethylation of the C28-sterol intermediate has been demonstrated. With 4α-methyl-5α-cholest-7-en-3β-ol as substrate, maximal velocities of ester formation (0.36 nmole/30 min per mg of protein) were almost equivalent to maximal velocities of demethylation (0.45 nmole/30 min per mg of protein). Ester formation may be eliminated by carrying out incubations with microsomal preparations; ester formation may be restored completely upon addition (to the microsomes) of either coenzyme A and ATP or the supernatant fraction resulting from centrifugation at 105,000 g.Ester formation has been examined similarly with broken-cell preparations of rat skin. With 4α-methyl-5α-cholest-7-en-3β-ol as substrate, the rate of ester formation was more than six times the rate of methyl sterol demethylation. The very significant competition between esterification and demethylation of methyl sterol intermediates of skin suggests that sterol intermediates accumulate in rat skin because of the rapid formation of esters that may not be further metabolized.
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