Nuclear protein that binds sterol regulatory element of low density lipoprotein receptor promoter. II. Purification and characterization

Xiaodong Wang, Michael R. Briggs, Xianxin Hua, Chieko Yokoyama, Joseph L. Goldstein, Michael S. Brown

Research output: Contribution to journalArticle

246 Scopus citations

Abstract

This paper describes the purification and characterization of a sterol regulatory element binding protein (SREBP) that recognizes the SRE-1 sequence in the 5' flanking region of the gene for the low density lipoprotein (LDL) receptor. The protein was purified more than 38,000-fold from nuclear extracts of human HeLa cells by ion exchange, gel filtration, and DNA- affinity chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified preparation revealed a cluster of bands at 59-68 kDa, each of which bound to the SRE-1 element as revealed by cross- linking experiments. Binding of SREBP correlated perfectly with transcriptional activity in a series of 16 sterol regulatory elements with point mutations. In the LDL receptor promoter, the 10-base pair SRE-1 is embedded in a 16-base pair sequence designated Repeat 2, which is adjacent to Repeat 3, a binding site for nuclear factor Sp1. Oligonucleotides containing Repeat 2 + 3 bound SREBP and Sp1 as revealed by mobility shift assays. SREBP produced a DNase I footprint over the SRE-1 sequence, which was immediately adjacent to the footprint produced by Sp1. The current data are consistent with the concept that SREBP acts in concert with Sp1 to achieve high level, sterol-suppressible transcription of the gene for the LDL receptor.

Original languageEnglish (US)
Pages (from-to)14497-14504
Number of pages8
JournalJournal of Biological Chemistry
Volume268
Issue number19
StatePublished - Jan 1 1993

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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