Individual metal ligands play distinct functional roles in the zinc sensor Staphylococcus aureus CzrA.
J Mol Biol
; 356(5): 1124-36, 2006 Mar 10.
Article
em En
| MEDLINE
| ID: mdl-16406068
ABSTRACT
Recent studies on metalloregulatory proteins suggest that coordination number/geometry and metal ion availability in a host cytosol are key determinants for biological specificity. Here, we investigate the contribution that individual metal ligands of the alpha5 sensing site of Staphylococcus aureus CzrA (Asp84, His86, His97', and His100') make to in vitro metal ion binding affinity, coordination geometry, and allosteric negative regulation of DNA operator/promoter region binding. All ligand substitution mutants exhibit significantly reduced metal ion binding affinity (K(Me)) by > or =10(3) M(-1). Substitutions of Asp84 and His97 give rise to non-native coordination geometries upon metal binding and are non-functional in allosteric coupling of metal and DNA binding (DeltaG(coupling) approximately 0 kcal mol(-1)). In contrast, His86 and His100 could be readily substituted with potentially liganding (Asp, Glu) and poorly liganding (Asn, Gln) residues with significant native-like tetrahedral metal coordination geometry retained in these mutants, leading to strong functional coupling (DeltaG(coupling) > or = +3.0 kcal mol(-1)). 1H-(15)N heteronuclear single quantum coherence (HSQC) spectra of wild-type and mutant CzrAs reveal that all H86 and H100 substitution mutants undergo 4 degrees structural switching on binding Zn(II), while D84N, H97N and H97D CzrAs do not. Thus, only those variant CzrAs that retain some tetrahedral coordination geometry characteristic of wild-type CzrA upon metal binding are capable of driving 4 degrees structural conformational changes linked to allosteric regulation of DNA binding in vitro, irrespective of the magnitude of K(Me).
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Staphylococcus aureus
/
Proteínas de Bactérias
/
Zinco
/
Isoformas de Proteínas
/
Proteínas de Ligação a DNA
/
Metais
Tipo de estudo:
Prognostic_studies
Idioma:
En
Revista:
J Mol Biol
Ano de publicação:
2006
Tipo de documento:
Article
País de afiliação:
Estados Unidos