Identification of putative two-component regulatory systems in the bovine pathogen Mannheimia haemolytica A1, and preliminary characterization of the NarQ/P system

Abstract The complete genome sequence of the bovine pathogen Mannheimia haemolytica A1 was analyzed by blast searches for the presence of two-component regulatory system proteins. Five complete sets of putative two-component systems were identified, and the NarQ/P system was further investigated. in silico analysis of the NarQ and NarP proteins showed features that are typical of the sensor and response regulator proteins. A narP knock-out mutant was constructed. The narP mutant has lost its ability to respond to NaNO3 in the media and fail to alter the expression of several proteins. One of the proteins that showed increased production in the parent strain in response to NaNO3 was analyzed by matrix-assisted laser desorption ionization time-of-flight MS. Unexpectedly, the protein was identified to be FbpA, a periplasmic component of the iron transporter system. Sequence analysis of the promoter region of fbpA identified motifs typical for NarP-regulated genes. The expression of the leukotoxin gene was also altered in the narP mutant as shown by Western immunoblot analysis and reverse transcription-PCR.