title: Cloning of the gene encoding DNA binding protein HU from Bacillus stearothermophilus and its expression in Escherichia coli.
abstract: The gene (hbst) encoding the DNA binding protein HU from Bacillus stearothermophilus was cloned, with the Bacillus subtilis HU gene (hbsu) as a hybridization probe. The nucleotide sequence, which contains a ribosome binding site, a transcriptional termination signal, as well as the coding region, was analyzed by the dideoxy chain-termination method. The deduced amino acid sequence of an open reading frame was perfectly matched with that of B. stearothermophilus HU (BstHU) determined by the protein chemical methods [M. Kimura and K. S. Wilson, J. Biol. Chem., 258, 4007-4011 (1983)]. The gene, hbst, was overexpressed using the expression vector pET-5a in Escherichia coli, and the recombinant HU protein (r-BstHU) was purified to be homogeneity by heparin-agarose column chromatography followed by ion-exchange column chromatography on S-Sepharose. The recombinant protein thus obtained had a circular dichroism spectrum identical to that of the authentic protein and bound to DNA to the same extent as the authentic protein.
authors.affiliation: Laboratory of Biochemistry, Faculty of Agriculture, Kyushu University, Fukuoka, Japan.
authors.forename: S
authors.initials: S
authors.lastname: Kawamura
authors.forename: H
authors.initials: H
authors.lastname: Kajiyama
authors.forename: N
authors.initials: N
authors.lastname: Yamasaki
authors.forename: M
authors.initials: M
authors.lastname: Kimura
mesh_terms: D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000235:genetics* / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D001483:Base Sequence; D015139:Blotting, Southern; D002852:Chromatography, Ion Exchange; D002942:Circular Dichroism; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics; D001411:Geobacillus stearothermophilus / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
chemical_list: D001426:Bacterial Proteins; D017931:DNA Primers; D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; C040330:histone-like protein HU, bacteria