HTC1, A SALICYLATE CHEMORECEPTOR FROM HALOMONAS TITANICAE KHS3

HERRERA SEITZ, M.K.; JUNG, K; PROSA, L.M.; STUDDERT, C. A.; GASPEROTTI, A. F.; BALMACEDA, R.S.

New Orleans

Congreso; BLAST (Bacterial locomotion and signal transduction) XIV Meeting; 2017

Background: Most chemoreceptors are dimeric transmembrane proteins with a periplasmic domain for ligand binding and a very conserved cytoplasmic domain for kinase control. Halomonas titanicae KHS3 was isolated from Mar del Plata harbor (Argentina), and has shown chemotaxis responses towards aromatic hydrocarbons. Two chemosensory clusters and 25 putative chemoreceptors were identified in its genomic sequence.Methods: Chemotaxis ability of H. titanicae KHS3 to different substrates was assessed in soft agar plates. Chemoreceptors were identified in H. titanicae genomic sequence based on the annotation obtained from RAST and IMG servers. Both the entire sequence and predicted periplasmic domain of H. titanicae chemoreceptor 1 (Htc1) were cloned and expressed in E. coli strains. The periplasmic domain was purified and subjected to thermal shift and thermophoresis assays in the presence of different substances. Flagellar control assays were performed in E. coli cells expressing Htc1 as the only receptor. Results: H. titanicae KHS3 displays chemotaxis both towards phenanthrene and sodium salicylate in swimming plates. Htc1 was found in a genomic context that suggested a role in aromatic compounds sensing. Thermal shift assays showed that the melting temperature of its purified periplasmic domain was increased by 4 degrees in the presence of 10 mM sodium salicylate, while it was unchanged in the presence of pyruvate, serine or phenanthrene. Thermophoresis assays carried out at different salicylate concentrations resulted in a Kd of 73 µM. E. coli cells expressing Htc1 as their only receptor responded to the addition of salicylate (but not phenanthrene) by suppressing CW rotation of their flagella. Conclusion: Htc1 is a chemoreceptor for sodium salicylate, based on its binding abilities and the flagellar control of E. coli cells. Even though Htc1 does not bind phenanthrene, it could be responsible of chemotaxis towards aromatic hydrocarbons, considering that salicylate is an intermediate metabolite of aromatic compounds degradation pathways. Gene replacement experiments to answer this question are in progress.