Department

Chemistry & Biochemistry

Document Type

Article

Publication Date

3-23-2015

Abstract

Flagellar biogenesis is a complex process that involves multiple checkpoints to coordinate transcription of flagellar genes with the assembly of the flagellum. In Helicobacter pylori, transcription of the genes needed in the middle stage of flagellar biogenesis is governed by RpoN and the two-component system consisting of the histidine kinase FlgS and response regulator FlgR. In response to an unknown signal, FlgS autophosphorylates and transfers the phosphate to FlgR, initiating transcription from RpoNdependent promoters. In the present study, export apparatus protein FlhA was examined as a potential signal protein. Deletion of its N-terminal cytoplasmic sequence dramatically decreased expression of two RpoN-dependent genes, flaB and flgE. Optical biosensing demonstrated a high-affinity interaction between FlgS and a peptide consisting of residues 1 to 25 of FlhA (FlhANT). The KD (equilibrium dissociation constant) was 21 nM and was characterized by fast-on (kon 2.9 104 M 1 s 1 ) and slow-off (koff 6.2 10 4 s 1 ) kinetics. FlgS did not bind peptides consisting of smaller fragments of the FlhANT sequence. Analysis of binding to purified fragments of FlgS demonstrated that the C-terminal portion of the protein containing the kinase domain binds FlhANT. FlhANT binding did not stimulate FlgS autophosphorylation in vitro, suggesting that FlhA facilitates interactions between FlgS and other structures required to stimulate autophosphorylation.

Journal

Journal of Bacteriology

Volume

197

Issue

11

First Page

1886

Last Page

1892

Digital Object Identifier (DOI)

10.1128/JB.02610-14

Included in

Chemistry Commons

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