Structural Insights into Homo- and Heterotropic Allosteric Coupling in the Zinc Sensor S. Aureus Czra from Covalently Fused Dimers.
From: Department of Biochemistry and Biophysics, Texas A&M University, College Station, TX 77843-2128, USA.
Journal of the American Chemical Society
- Publish Date: Feb 2006
- ISSN: 0002-7863
- Volume: 128
- Issue: 6
- Pages: 1937-47
- Medium: Print
- Language: English
- Citation (JAMA): Lee Sunbae, Arunkumar Alphonse I, Chen Xiaohua, et al. Structural Insights into Homo- and Heterotropic Allosteric Coupling in the Zinc Sensor S. Aureus Czra from Covalently Fused Dimers.. J. Am. Chem. Soc. Feb 2006;128:1937-47
Abstract
The Zn(II)/Co(II)-sensing transcriptional repressor, Staphylococcus aureus CzrA, is a homodimer containing a symmetry-related pair of subunit-bridging tetrahedral N(3)O metal sensor coordination sites. A metal-induced quaternary structural change within the homodimer is thought to govern the biological activity of this and other metal sensor proteins. Here, we exploit covalent (Gly(4)Ser)(n)() linkers of variable length in “fused” CzrAs, where n = 1 (designated 5L-fCzrA), 2 (10L-fCzrA), or 3 (15L-fCzrA), as molecular rulers designed to restrict any quaternary structural changes that are associated with metal binding and metal-mediated allosteric regulation of DNA binding to varying degrees. While 15L-fCzrA exhibits properties most like homodimeric CzrA, shortening the linker in 10L-fCzrA abolishes negative homotropic cooperativity of Zn(II) binding and reduces DNA binding affinity of the apoprotein significantly. Decreasing the linker length further in 5L-fCzrA effectively destroys one metal site altogether and further reduces DNA binding affinity. However, Zn(II) negatively regulates DNA binding of all fCzrAs, with allosteric coupling free energies (DeltaG(1)(c)) of 4.6, 3.1, and 2.7 kcal mol(-1) for 15L-, 10L-, and 5L-fCzrAs, respectively. Introduction of a single nonliganding H97N substitution into either the N-terminal or C-terminal protomer domain in 10L-fCzrA results in DeltaG(1)(c) = 2.6 kcal mol(-1) or approximately 83% that of 10L-fCzrA; in contrast, homodimeric H97N CzrA gives DeltaG(1)(c) = 0. (1)H-(15)N HSQC spectra acquired for wt-, 10L-fCzrA and H97N 10L-fCzrA in various Zn(II) ligation states reveal that the allosteric change of the protomer domains within the fused dimer is independent and not concerted. Thus, occupancy of a single metal site by Zn(II) introduces asymmetry into the CzrA homodimer that leads to significant allosteric regulation of DNA binding.
Mesh Headings (Keywords): Bacterial Proteins, Base Sequence, Binding Sites, DNA, DNA-Binding Proteins, Dimerization, Kinetics, Models, Molecular, Molecular Sequence Data, Structure-Activity Relationship, Thermodynamics, Zinc
Check for Full Text / PubMed Unique Identifier (PMID): 16464095
This abstract is part of PubMed, a service of the U.S. National Library of Medicine. PubMed includes more than 17 million citations from MEDLINE and other life science journals for biomedical articles. See Copyright and Disclaimers.
Linked medical terms appearing on this page are added by Healia to help readers find more information and are not part of the original PubMed document.
The data herein was last updated on July 8th, 2008 and may not reflect the most current and accurate data available from NLM.