title: Cobalt-substituted zinc finger 3 of transcription factor IIIA: interactions with cognate DNA detected by (31)P ENDOR spectroscopy.
abstract: We show the first ENDOR study of the coordination environment of high-spin Co(II) in a biological system with a study of DNA binding to the Co-substituted Cys2/His2 single Zn-finger domain, Finger 3 (F3), from the prototypical zinc finger protein, transcription factor IIIA (TFIIIA) from Xenopus laevis. High covalency to cysteine and histidine is implied by ENDOR-derived 1H couplings to protons of cysteinyl ligands and 14N couplings to histidyl nitrogens, results which support the expectation that Zn(II) and Co(II) bind to F3 in a very similar manner. No changes in either 1H or 14N ENDOR were detected upon binding Co(II)-F3 to C-block DNA. Of particular importance to the use of Co(II) substitution for Zn(II), the ENDOR method shows that Co(II)-F3 undergoes sequence-specific binding to the cognate DNA for Zn(II)-F3, the internal control region (ICR) of the 5S rRNA (C-block). 31P ENDOR measurements yield a Co-31P distance of rCo-P = 8.1(3) A to the nearest backbone phosphodiester of the C-block. Interestingly, a 31P ENDOR doublet observed for Co(II)-F3 in phosphate buffer indicates that inorganic phosphate (Pi) binds at a comparable distance from Co as does the nearest phosphate of DNA, presumably at the same site.
authors.affiliation: Department of Chemistry, Northwestern University, Evanston, IL 60208-3113, USA.
authors.forename: Charles J
authors.initials: CJ
authors.lastname: Walsby
authors.forename: Dmitriy
authors.initials: D
authors.lastname: Krepkiy
authors.forename: David H
authors.initials: DH
authors.lastname: Petering
authors.forename: Brian M
authors.initials: BM
authors.lastname: Hoffman
mesh_terms: D003035:Cobalt / Q000737:chemistry*; D004247:DNA / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods; D008958:Models, Molecular; D009841:Oligonucleotides / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet; D035582:Transcription Factor TFIIIA / Q000737:chemistry*; D016335:Zinc Fingers
chemical_list: D009841:Oligonucleotides; D035582:Transcription Factor TFIIIA; D003035:Cobalt; D004247:DNA