NOVEL TOPOLOGY OF A ZINC-BINDING DOMAIN FROM A PROTEIN INVOLVED IN REGULATING EARLY XENOPUS DEVELOPMENT

Xenopus nuclear factor XNF7, a maternally expressed protein, functions in patterning of the embryo, XNF7 contains a number of defined protein domains implicated in the regulation of some developmental processes. Among these is a tripartite motif comprising a zinc-binding RING finger and B-box domain next to a predicted alpha-helical coiled-coil domain, Interestingly, this motif is found in a variety of proteins including several proto-oncoproteins. Here we describe the solution structure of the XNF7 B-box zinc-binding domain determined at physiological pH by H-1 NMR methods, The B-box structure represents the first three-dimensional structure of this new motif and comprises a monomer having two beta-strands, two helical turns and three extended loop regions packed in a novel topology. The r.m.s. deviation for the best 18 structures is 1.15 Angstrom for backbone atoms and 1.94 Angstrom for all atoms, Structure calculations and biochemical data shows one zinc atom ligated in a Cys2-His2 tetrahedral arrangement. We have used mutant peptides to determine the metal ligation scheme which surprisingly shows that not all of the seven conserved cysteines/histidines in the B-box moth are involved in metal ligation. The B-box structure is not similar in tertiary fold to any other known zinc-binding motif.