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Effects of select histidine to cysteine mutations on transcriptional regulation by Escherichia coli RcnR.
- Source :
-
Biochemistry [Biochemistry] 2013 Jan 08; Vol. 52 (1), pp. 84-97. Date of Electronic Publication: 2012 Dec 24. - Publication Year :
- 2013
-
Abstract
- The RcnR metalloregulator represses the transcription of the Co(II) and Ni(II) exporter, RcnAB. Previous studies have shown that Co(II) and Ni(II) bind to RcnR in six-coordinate sites, resulting in derepression. Here, the roles of His60, His64, and His67 in specific metal recognition are examined. His60 and His64 correspond to ligands that are important for Cu(I) binding in the homologous Cu(I)-responsive metalloregulator, CsoR. These residues are known to be functionally important in RcnR transcriptional regulation. X-ray absorption spectroscopy (XAS) was used to examine the structure of bound cognate and noncognate metal ions, and lacZ reporter assays were used to assess the transcription of rcnA in response to metal binding in the three His → Cys mutations, H60C, H64C, and H67C. These studies confirm that both Ni(II) and Co(II) use His64 as a ligand. H64C-RcnR is also the only known mutant that retains a Co(II) response while eliminating the response to Ni(II) binding. XAS data indicate that His60 and His67 are potential Co(II) ligands. The effects of the mutations of His60, His64, and His67 on the structures of the noncognate metal ions [Zn(II) and Cu(I)] reveal that these residues have distinctive roles in binding noncognate metals. None of the His → Cys mutants in RcnR confer any response to Cu(I) binding, including H64C-RcnR, where the ligands involved in Cu(I) binding in CsoR are present. These data indicate that while the secondary, tertiary, and quaternary structures of CsoR and RcnR are quite similar, small changes in primary sequence reveal that the specific mechanisms involved in metal recognition are quite different.
- Subjects :
- Amino Acid Sequence
Cobalt metabolism
Cysteine metabolism
Escherichia coli chemistry
Escherichia coli metabolism
Escherichia coli Proteins chemistry
Escherichia coli Proteins metabolism
Histidine metabolism
Molecular Sequence Data
Nickel metabolism
Point Mutation
Protein Conformation
Repressor Proteins chemistry
Repressor Proteins metabolism
Sequence Alignment
Transcriptional Activation
X-Ray Absorption Spectroscopy
Cysteine genetics
Escherichia coli genetics
Escherichia coli Proteins genetics
Gene Expression Regulation, Bacterial
Histidine genetics
Membrane Proteins genetics
Repressor Proteins genetics
Subjects
Details
- Language :
- English
- ISSN :
- 1520-4995
- Volume :
- 52
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- Biochemistry
- Publication Type :
- Academic Journal
- Accession number :
- 23215580
- Full Text :
- https://doi.org/10.1021/bi300886q