Substrate specificity of Helicobacter pylori histone-like HU protein is determined by insufficient stabilization of DNA flexure points.
The histone-like HU protein is ubiquitous in the eubacteria. A role for Escherichia coli HU in compaction of the bacterial genome has been reported, along with regulatory roles in DNA replication, transposition, repair and transcription. We show here that HU from the human pathogen Helicobacter pylori, which has been implicated in the development of ulcers and gastric cancer, exhibits enhanced thermal stability and distinct DNA substrate specificity. Thermal denaturation of HpyHU (H. pylori HU) measured by CD spectroscopy yields a melting temperature (T(m)) of 56.4+/-0.1 degrees C. HpyHU binds linear duplex DNA with a site size of approximately 19 bp and with low affinity, but in striking contrast to E. coli HU, HpyHU has only modest preference for DNA with mismatches, nicks or gaps. Instead, HpyHU binds stably to four-way DNA junctions with half-maximal saturation of 5 nM. Substitution of two residues adjacent to the DNA-intercalating prolines attenuates both the preference for flexible DNA and the ability to bend and supercoil DNA. These observations suggest that proline intercalation generates hinges that must be stabilized by adjacent residues; insufficient stabilization leads to reduced bending and a failure to bind preferably to DNA with flexure points, such as gaps and mismatches.
Chen
Christina
C
Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA.
Ghosh
Sharmistha
S
Grove
Anne
A
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004247:DNA / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D016480:Helicobacter pylori / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D006657:Histones / Q000737:chemistry*; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010957:Plasmids / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D013379:Substrate Specificity
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D006657:Histones; D004247:DNA