19F-NMR reveals metal and operator-induced allostery in MerR.
Metalloregulators of the MerR family activate transcription upon metal binding by underwinding the operator-promoter DNA to permit open complex formation by pre-bound RNA polymerase. Historically, MerR's allostery has been monitored only indirectly via nuclease sensitivity or by fluorescent nucleotide probes and was very specific for Hg(II), although purified MerR binds several thiophilic metals. To observe directly MerR's ligand-induced behavior we made 2-fluorotyrosine-substituted MerR and found similar, minor changes in (19)F chemical shifts of tyrosine residues in the free protein exposed to Hg(II), Cd(II) or Zn(II). However, DNA binding elicits large chemical shift changes in MerR's tyrosine residues and in DNA-bound MerR Hg(II) provokes changes very distinct from those of Cd(II) or Zn(II). These chemical shift changes and other biophysical and phenotypic properties of wild-type MerR and relevant mutants reveal elements of an allosteric network that enables the coordination state of the metal binding site to direct metal-specific movements in the distant DNA binding site and the DNA-bound state also to affect the metal binding domain.
Song
Lingyun
L
Department of Microbiology, University of Georgia, Athens, GA 30602, USA.
Teng
Quincy
Q
Phillips
Robert S
RS
Brewer
John M
JM
Summers
Anne O
AO
D000494:Allosteric Regulation; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D002104:Cadmium / Q000737:chemistry / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005462:Fluorine Radioisotopes / Q000737:chemistry / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D008628:Mercury / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019906:Nuclear Magnetic Resonance, Biomolecular; D017434:Protein Structure, Tertiary; D012045:Regulatory Sequences, Nucleic Acid; D016415:Sequence Alignment; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D015032:Zinc / Q000737:chemistry / Q000378:metabolism
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D005462:Fluorine Radioisotopes; C051124:MerR protein, Bacteria; D002104:Cadmium; D014443:Tyrosine; C060489:2-fluorotyrosine; D008628:Mercury; D015032:Zinc