Cholesterol biosynthesis from lanosterol: regulation and purification of rat hepatic sterol 14-reductase

Chap Ki Kim, Kye Im Jeon, Dong Min Lim, Tae Neung Johng, James M. Trzaskos, James L. Gaylor, Young Ki Paik

Research output: Contribution to journalArticle

39 Citations (Scopus)

Abstract

We have previously characterized the membrane-bound sterol 14-reductase (14-reductase) that catalyzes anaerobically NADPH-dependent reduction of the 14-double bond of Δ8,14-diene or Δ7,14-diene sterols that are sterol intermediates in cholesterol biosynthesis mammals (Paik et al. (1984) J. Biol. Chem. 259, ]3413-13423). To elucidate the regulatory mechanism as well as molecular characteristics of the 14-reductase, we extended our investigation on the consequences of alteration of the enzymic activity under various physiological conditions. The enzymic activity of rat hepatic sterol 14-reductase was induced more than 11-fold by feeding 5% cholestyramine plus 0.1 % lovastatin (the CL-diet) for 7 days but was severely suppressed by feeding 5% cholesterol or 0.01 % AY-9944 (an inhibitor of 14-reductase) for the same period. The increase or decrease in the 14-reductase activity also parallels the same change in the cholesterol synthetic rate in hepatocytes from rats that had been fed either the CL-diet or 0.01 % AY-9944. In vitro inhibition studies revealed that AY-9944 acts as a competitive inhibitor of the 14-reductase (Ki = 0.26 μM). A diurnal variation was observed for the 14-reductase with peak activity near the middle of the dark cycle (10 p.m.), which was abolished by administration of cycloheximide. With induced enzyme conditions 14-reductase has been further purified with chromatographic procedures to near homogeneity. Purified 14-reductase appears to be a Mr = 70 000 protein that is composed of two equally-sized subunits having a Mr = 38 000. All properties of the purified 14-reductase suggest that the solubilized enzyme is the principal 14-reductase of microsomes. Taken together, our results provide the first evidence in support of a previously unknown regulatory role for the 14-reductase in the overall cholesterol synthetic pathway.

Original languageEnglish
Pages (from-to)39-48
Number of pages10
JournalBiochimica et Biophysica Acta (BBA)/Lipids and Lipid Metabolism
Volume1259
Issue number1
DOIs
Publication statusPublished - 1995 Oct 26

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Endocrinology

Fingerprint Dive into the research topics of 'Cholesterol biosynthesis from lanosterol: regulation and purification of rat hepatic sterol 14-reductase'. Together they form a unique fingerprint.

  • Cite this