title: The six zinc fingers of metal-responsive element binding transcription factor-1 form stable and quasi-ordered structures with relatively small differences in zinc affinities.
abstract: Six Cys(2)His(2) zinc fingers (F1-6) comprise the DNA binding domain of metal-responsive element binding transcription factor-1 (MTF-1). F1-6 is necessary for basal and zinc-induced expression of metallothionein genes. Analysis of NMR structural and dynamic data for an F1-6 protein construct demonstrates that each zinc finger adopts a stable betabetaalpha fold in the presence of stoichiometric Zn(II), provided that all cysteine ligands are in a reduced state. Parallel studies of protein constructs spanning the four N-terminal core DNA binding fingers (F1-4) and two C-terminal low DNA affinity fingers (F5-6) reveal similar stable zinc finger structures. In both the F1-6 and F5-6 proteins, the finger 5 cysteines were found to readily oxidize at neutral pH. Detailed spectral density and hydrodynamic analysis of (15)N relaxation data revealed quasi-ordered anisotropic rotational diffusion properties of the six F1-6 zinc fingers that could influence MTF-1 DNA binding function. A more general effect on the rotational diffusion properties of Cys(2)His(2) zinc fingers was also uncovered that is dependent upon the position of each finger within multifinger domains. Analysis of NMR (1)H-(15)N-heteronuclear single quantum coherence spectral peak intensities measured as a function of added Zn(II) in conjunction with Zn(II) binding modeling studies indicated that the Zn(II) affinities of all MTF-1 zinc fingers are within approximately 10-50-fold. These analyses further suggested that metal sensing by MTF-1 in eukaryotic cells involves multiple zinc fingers and occurs over a 100-fold or less range of accessible Zn(II) concentration.
authors.affiliation: Division of Cell Biology and Biophysics, School of Biological Sciences, University of Missouri-Kansas City, Kansas City, Missouri 64110, USA.
authors.forename: Belinda M
authors.initials: BM
authors.lastname: Potter
authors.forename: Linda S
authors.initials: LS
authors.lastname: Feng
authors.forename: Priya
authors.initials: P
authors.lastname: Parasuram
authors.forename: Viktor A
authors.initials: VA
authors.lastname: Matskevich
authors.forename: Jed A
authors.initials: JA
authors.lastname: Wilson
authors.forename: Glen K
authors.initials: GK
authors.lastname: Andrews
authors.forename: John H
authors.initials: JH
authors.lastname: Laity
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D004268:DNA-Binding Proteins; D007700:Kinetics; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015032:Zinc / Q000378:metabolism*; D016335:Zinc Fingers; D000097809:Transcription Factor MTF-1
chemical_list: D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D014157:Transcription Factors; D015032:Zinc; D000097809:Transcription Factor MTF-1