Domains of Mnt repressor: roles in tetramer formation, protein stability, and operator DNA binding

Abstract

The Mnt repressor of bacteriophage P22 is a member of the ribbon-helix-helix family of gene regulatory proteins. Proteolytic cleavage of Mnt with chymotrypsin reveals that it consists of two structural domains. Both domains are required for high-affinity operator binding. The N domain (residues 1-51) is dimeric and binds weakly but specifically to operator DNA. The C domain (residues 52-82) forms an independent alpha-helical, tetramerization domain and, by itself, has no DNA-binding activity. In intact Mnt, the N and C domains help to stabilize each other against denaturation but appear to be linked rather flexibly. Assays of the half-operator affinities of Mnt and the isolated N domain indicate that binding to adjacent half-sites in the whole operator is stabilized by protein-protein contacts between N domains in addition to protein-protein contacts between C domains.