MODULATION OF DNA-BINDING OF A TRAMTRACK ZINC-FINGER PEPTIDE BY THE METALLOTHIONEIN-THIONEIN CONJUGATE PAIR

The ability of metallothionein (MT) to modulate DNA binding by a two-f inger peptide of Tramtrack (TTK), a CCHH zinc transcription factor, wa s investigated using metal bound and metal-deficient forms of rabbit M T-2 and the TTK peptide. Thionein inhibited DNA binding by zinc-bound TTK, and Zn-MT restored DNA-binding by zinc deficient apo-TTK. ''Free' ' zinc at low concentrations was as effective as Zn-MT in restoring DN A binding by apopeptide but was inhibitory at concentrations equal to zinc bound to 2 mol eq and higher of Zn-MT. Substitution of cadmium fo r zinc reduced the affinity of the peptide for its DNA binding site. T his effect was reversed by incubation with Zn-MT. The circular dichroi c spectra of the TTK peptide indicated that zinc removal resulted in l oss of a-helical structures, which are sites of DNA contact points. Re constitution with cadmium resulted in stoichiometric substitution of 2 mol of Cd/mol of peptide but not recovery of a-helical structures. In cubation of Cd-TTK with Zn MT restored the secondary structure expecte d for zinc-bound TTK. The ability of Zn-MT and thionein to restore or inhibit DNA-binding by TTK was associated with effects on the metallat ion status of the peptide and related alterations in its secondary str ucture.