title: Purification and functional analysis of recombinant Acholeplasma laidlawii histone-like HU protein.
abstract: HU is a most abundant DNA-binding protein in bacteria. This protein is conserved either in its heterodimeric form or in one of its homodimeric forms in all bacteria, in plant chloroplasts, and in some viruses. HU protein non-specifically binds and bends DNA as a hetero- or homodimer and can participate in DNA supercoiling and DNA condensation. It also takes part in some DNA functions such as replication, recombination, and repair. HU does not recognize any specific sequences but shows some specificity to cruciform DNA and to repair intermediates, e.g., nick, gap, bulge, 3'-overhang, etc. To understand the features of HU binding to DNA and repair intermediates, a fast and easy HU proteins purification procedure is required. Here we report overproduction and purification of the HU homodimers. The method of HU purification allows obtaining a pure recombinant non-tagged protein cloned in Escherichia coli. We applied this method for purification of Acholeplasma laidlawii HU and demonstrated that this protein possesses a DNA-binding activity and is free of contaminating nuclease activity. Besides that we have shown that expression of A. laidlawii ihf_hu gene in a slow-growing hupAB E. coli strain restores the wild-type growth indicating that aclHU can perform the basic functions of E. coli HU in vivo.
authors: Research Institute for Physico-Chemical Medicine of Federal Medical-Biological Agency of Russian Federation, Malaya Pirogovskaya str. 1A, Moscow, Russia. krolick@yandex.ru
authors: S A
authors: SA
authors: Levitskiy
authors: A M
authors: AM
authors: Sycheva
authors: D D
authors: DD
authors: Kharlampieva
authors: J
authors: J
authors: Oberto
authors: D E
authors: DE
authors: Kamashev
authors: M V
authors: MV
authors: Serebryakova
authors: S A
authors: SA
authors: Moshkovskii
authors: V N
authors: VN
authors: Lazarev
authors: V M
authors: VM
authors: Govorun
mesh_terms: D000128:Acholeplasma laidlawii / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004270:DNA, Circular / Q000235:genetics / Q000378:metabolism; D004278:DNA, Superhelical / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development; D005816:Genetic Complementation Test; D009154:Mutation; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D055503:Protein Multimerization; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D020349:Surface Plasmon Resonance
chemical_list: D001426:Bacterial Proteins; D004270:DNA, Circular; D004278:DNA, Superhelical; D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D004247:DNA