HU-1 MUTANTS OF ESCHERICHIA-COLI DEFICIENT IN DNA-BINDING

We constructed four mutants of the Escherichia coli hupB gene, encoding HU-1 protein, by synthetic oligodeoxyribonucleicotide-directed, site-specific mutagenesis on M13mp18 vectors. The HupBR45 protein contained alterations of Arg58 → Gly and Arg61 → Gly, and the HupBF3, HupBK2 and HupBK2 and HupBA1 proteins contained Phe47 → Thr, Lys37 → Gln and Ala30 → Asp alterations, respectively. HupBF3 and HupBR45 were unable to maintain normal cell growth in a hupA-hupB-himA triple mutant at 42°C, mini-F or RSF1010 proliferation, or Mu phage development in a hupA-hupB double mutant, whereas HupBA1 and HupBK2 supported these cellular activities. DNA-affinity column chromatography showed that the HupBF3 and HupBR45 had reduced affinities to DNA. These observations indicate that two highly conserved Arg residues in the arm structure of the C-terminal half of the HU-1 molecule and a Phe residue in the short β-sheet connecting the two halves of the molecule are important for the DNA-binding ability and biological functions of this protein. © 1990.