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PMID: 7630724 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S.

Metalloregulation of the cyanobacterial smt locus: identification of SmtB binding sites and direct interaction with metals.

Nucleic acids research ·Vol. 23 ·No. 13 ·1995-07-11 ·Pages 2472-8

Erbe JL, Taylor KB, Hall LM

Abstract

The smtB gene of Synechococcus PCC 7942 encodes a trans-acting repressor of the metal-regulated smtA gene that encodes a class II metallothionein. Recombinant SmtB has been expressed in Escherichia coli and purified. Electrophoretic mobility shift assays using recombinant SmtB or a protein extract from Synechococcus PCC 6301 reveal the concentration-dependent formation of three specific complexes with the smt operator/promoter. SmtB is also capable of direct interaction with metals as evidenced by 65Zn binding to the SmtB protein as well as the inhibition of repressor-DNA complex formation in the presence of various metal ions. Methylation interference analysis of such complexes identifies four protein contact points within the smt operator/promoter DNA. The points of contact appear to represent two pairs of binding sites, one pair in each of two inverted repeats (nt 548-563, 589-602). The contact points within each pair lie on opposing DNA strands and are separated by 10 bp, placing the repressor binding sites on opposite sides of the DNA helix. Based on electrophoretic mobility shift assays, methylation interference and molecular size calculations we propose that recombinant SmtB binds to the smt operator/promoter in multimeric fashion.

Related Genes
smtB
MeSH Terms
Bacterial Proteins Base Sequence Binding Sites Cyanobacteria/genetics DNA, Bacterial/chemistry,metabolism DNA-Binding Proteins/genetics,metabolism Edetic Acid/pharmacology Gene Expression Regulation/drug effects Metals/pharmacology Molecular Sequence Data Operon Promoter Regions, Genetic Recombinant Proteins/metabolism Repressor Proteins/genetics,metabolism Zinc/pharmacology Zinc Radioisotopes
Chemicals
Bacterial Proteins DNA, Bacterial DNA-Binding Proteins Metals Recombinant Proteins Repressor Proteins Zinc Radioisotopes Edetic Acid Zinc
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Erbe J L
Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham 35294, USA.
Taylor K B
Hall L M
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
0305-1048
DOI
10.1093/nar/23.13.2472
Published
1995-07-11
Pages
2472-8
Language
English
Region
England
NLM ID
0411011
PMCID
PMC307053
Subset
IM
External Links
PubMed source DOI: 10.1093/nar/23.13.2472 PMC Full Text View journal details
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