Helicobacter pylori (H. pylori) colonizes under harsh acidic/oxidative stress conditions of human gastrointestinal tract and can survive there for infinitely longer durations of host life. The bacterium expresses several harbinger proteins to facilitate its persistent colonization under such conditions. One such protein in H. pylori is histone-like DNA binding protein (Hup), which in its homo-dimeric form binds to DNA to perform various DNA dependent cellular activities. Further, it also plays an important role in protecting the genomic DNA from oxidative stress and acidic denaturation. Legitimately, if the binding of Hup to DNA is suppressed, it will directly impact on the survival of the bacterium, thus making Hup a potential therapeutic target for developing new anti-H. pylori agents. However, to inhibit the binding of Hup to DNA, it is necessary to gain detailed insights into the molecular and structural basis of Hup-dimerization and its binding mechanism to DNA. As a first step in this direction, we report here the nuclear magnetic resonance (NMR) assignments and structural features of Hup at pH 6.0. The study revealed the occurrence of dynamic equilibrium between its monomer and dimer conformations. The dynamic equilibrium was found to shifting towards dimer both at low temperature and low pH; whereas DNA binding studies evidenced that the protein binds to DNA in its dimeric form. These preliminary investigations correlate very well with the diverse functionality of protein and will form the basis for future studies aiming to develop novel anti-H. pylori agents employing structure-based-rational drug discovery approach.
The summarized amalgam of internal
relaxation modulations and external
forces like pH, temperature, and solvent conditions determine the
protein structure, stability, and function. In a free-energy landscape,
although conformers are arranged in vertical hierarchy, there exist
several adjacent parallel sets with conformers occupying equivalent
energy cleft. Such conformational states are pre-requisites for the
functioning of proteins that have oscillating environmental conditions.
As these conformational changes have utterly small re-arrangements,
nuclear magnetic resonance (NMR) spectroscopy is unique in elucidating
the structure–dynamics–stability–function relationships
for such conformations. Helicobacter pylori survives and causes gastric cancer at extremely low pH also. However,
least is known as to how the genome of the pathogen is protected from
reactive oxygen species (ROS) scavenging in the gut at low pH under
acidic stress. In the current study, biophysical characteristics of H. pylori DNA binding protein (Hup) have been elucidated
at pH 2 using a combination of circular dichroism, fluorescence, NMR
spectroscopy, and molecular dynamics simulations. Interestingly, the
protein was found to have conserved structural features, differential
backbone dynamics, enhanced stability, and DNA binding ability at
low pH as well. In summary, the study suggests the partaking of Hup
protein even at low pH in DNA protection for maintaining the genome
integrity.
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