Nucleic Acids Research, 1993, Vol. 21, No. 7 1549-1554
© 1993
MOLECULAR BIOLOGY |
Purification of mouse MEP-1, a nuclear protein which binds to the metal regulatory elements of genes encoding metallothionein
Centre de Recherche en Cancérologie de l'Université Laval L'Hôtel-Dieu de Québec, 11 côte du Palais, Québec G1R 2J6 and Département de Physiologie, Facultié de Médecine, Université Laval Québec G1K 7P4, Canada
* To whom correspondence should be addressed
Received January 27, 1993. Accepted February 18, 1993.
Metal regulatory elements (MREs) shared by metallothionein (MT) gene promoters are essential for metal induction of MT genes. MEP-1, a nuclear protein which binds to these elements has been purified from heavy metal-resistant mouse L cells using footprinting, Southwestern and UV cross-linking techniques to assay its binding activity. The purification scheme, starting from crude nuclear extracts, involved a combination of heparin-Sepharose and MRE-DNA affinity chromatography. The purified protein preparation showed a single polypeptide band of 108 kDa on polyacrylamide gel electrophoresis, and 2D-gel analyses revealed the presence of a protein species migrating as a single population of approximately 110 kDa. MEP-1 does not appear to be glycosylated since it eluted with the flow-through on a Wheat Germ Sepharose column. It was retained by a zinc-Chelating Sepharose column suggesting that amino acid residues (i.e., cysteine, histidine) which have an affinity for zinc ions are exposed on the protein surface. Binding studies with the purified protein indicated that it binds specifically to MRE sequences and that the binding can be abolished by a point mutation in the MRE core consensus sequence or by the addition of the chelating agent 1,10-phenanthroline. Binding activity can be restored by the addition of zinc ions to the chelated protein. These results suggest that MEP-1 is one of the major proteins interacting with MRE sequences.
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  O. LaRochelle, S. Labbe, J.-F. Harrisson, C. Simard, V. Tremblay, G. St-Gelais, M. V. Govindan, and C. Seguin Nuclear Factor-1 and Metal Transcription Factor-1 Synergistically Activate the Mouse Metallothionein-1 Gene in Response to Metal Ions J. Biol. Chem., March 28, 2008; 283(13): 8190 - 8201. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. E. Hake, R. Mendez, and J. D. Richter Specificity of RNA Binding by CPEB: Requirement for RNA Recognition Motifs and a Novel Zinc Finger Mol. Cell. Biol., February 1, 1998; 18(2): 685 - 693. [Abstract] [Full Text]
|
|