PURIFICATION OF THE INTEGRATION HOST FACTOR HOMOLOG OF RHODOBACTER-CAPSULATUS - CLONING AND SEQUENCING OF THE HIP GENE, WHICH ENCODES THE BETA-SUBUNIT

We describe a method for rapid purification of the integration host fa ctor (IHF) homolog of Rhodobacter capsulatus that has allowed us to ob tain microgram quantities of highly purified protein. R. capsulatus IH F is an alphabeta heterodimer similar to IHF of Escherichia coli. We h ave cloned and sequenced the hip gene, which encodes the beta subunit. The deduced amino acid sequence (10.7 kDa) has 46% identity with the beta subunit of IHF from E. coli. In gel electrophoretic mobility shif t DNA binding assays, R. capsulatus IHF was able to form a stable comp lex in a site-specific manner with a DNA fragment isolated from the pr omoter of the structural hupSL operon, which contains the IHF-binding site. The mutated IHF protein isolated from the Hup- mutant IR4, which is mutated in the himA gene (coding for the alpha subunit), gave a sh ifted band of greater mobility, and DNase I footprinting analysis has shown that the mutated IHF interacts with the DNA fragment from the hu pSL promoter region differently from the way that the wild-type IHF do es.