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Publication Abstract from PubMed

Mycobacterium tuberculosis (Mtb) DosS is critical for the induction of Mtb dormancy genes in response to nitric oxide (NO), carbon monoxide (CO) or hypoxia. These environmental stimuli, which are sensed by the DosS heme group, result in autophosphorylation of a DosS His residue, followed by phosphotransfer to an Asp residue of the response regulator DosR. To clarify the mechanism of gaseous ligand recognition and signaling, we investigated the hydrogen-bonding interactions of the iron-bound CO and NO ligands by site-directed mutagenesis of Glu87 and His89. Autophosphorylation assays and molecular dynamics simulations suggest that Glu87 has an important role in ligand recognition, while His89 is essential for signal transduction to the kinase domain, a process for which Arg204 is important. Mutation of Glu87 to Ala or Gly rendered the protein constitutively active as a kinase, but with lower autophosphorylation activity than the wild-type in the Fe(II) and the Fe(II)-CO states, whereas the E87D mutant had little kinase activity except for the Fe(II)-NO complex. The H89R mutant exhibited attenuated autophosphorylation activity, while the H89A and R204A mutants were inactive as kinases, emphasizing the importance of these residues in communication to the kinase core. Resonance Raman of the wild-type and H89A mutant indicate the mutation does not alter the heme coordination number, spin state, or porphyrin deformation state, but suggests that interdomain interactions are disrupted by the mutation. Overall, these results confirm the importance of the distal hydrogenbonding network in ligand recognition and communication to the kinase domain, and reveal the sensitivity of the system to subtle differences in the binding of gaseous ligands.

Distal Hydrogen-Bonding Interactions in Ligand Sensing and Signaling by Mycobacterium tuberculosis DosS.,Basudhar D, Madrona Y, Yukl ET, Sivaramakrishnan S, Nishida CR, Moenne-Loccoz P, Ortiz de Montellano PR J Biol Chem. 2016 May 27. pii: jbc.M116.724815. PMID:27235395^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.