DNA RECOGNITION BY ZN[CYSTEINYL-HIS-OME](2) - A MINIMAL ZINC-FINGER DOCKING UNIT

Zinc fingers, 30-residue peptides anchored on Zn(II) coordinated to pa irs of cysteines and histidines, recognize DNA triplets and, as tandem modules, effect sequence read out. The focus of zinc finger-DNA inter action studies thus far has been to probe the nature of the binding of the 12-residue recognition element of the finger with DNA code bases. To understand the possible role of the Zn(II) ligand and to assess it s own DNA interaction profile, [(CH)(2)Zn] (C:cysteine, H:histidine; F igure 1) was constructed from bis-(t)Boc-cystinyl-di-His-OMe via thiol -disulfide exchange, Zn(II) complexation, and deprotection. [(CH)(2)Zn ] binds with polyd-(G . C). polyd(G . C) with association constants-1. 8x10(7) M(-1) (specific DNA-phosphate) and 3.3 x 10(3) M(-1) (nonspeci fic DNA-phosphate); perturbs its B-DNA profile; and enhances the T-m f rom 62.5 to 70.15 degrees C in a concentration-independent manner, wit h an ideal reversal profile on cooling, not observed in the DNA alone; releases polyd(G . C). polyd(G . C)-bound ethidium bromide; enhances the fluorescence of polyd(G . C). polyd(G . C)-bound ethidium bromide at low concentrations; and quenches it at higher ranges. [(CH)(2)ZN] a lso binds to d(ACGCTGGGCGT), the sequence associated with Zif-268, 3-f inger binding site. Such interactions were not seen in parallel studie s with (a) polyd(A . T). polyd(A . T) and [(CH)(2)Zn] and (b) {[C'H-2] (C':cystine; H:histidine; the direct metal-free precursor of [(CH)(2) Zn]}, ionic zinc nitrate, and covalent zinc acetyl acetonate Zn(AcAc)( 2), with poly[d(G . C). polyd(G . C)]. The results are rationalized on the basis of two types of association between [(CH)(2)Zn] and polyd(G . C). polyd(G . C), a nonspecific recognition of the sugar phosphate backbone, by an imidazole of [(CH)(2)Zn] and a specific one involving the amino group of [(CH)(2)Zn] and the guanine base of DNA. Control ex periments show that the latter greatly promotes DNA recognition. The p ossibility for such specific interactions with relatively small struct ures of the type [(CH)(2)Zn] would be of use in the design of DNA reco gnition elements and also provide an explanation for the experimentall y found variation in the placement of the zinc docking unit around the major groove of DNA. (C) 1997 John Wiley & Sons, Inc.