Identification and localization of a methyl-accepting chemotaxis protein in Rhodobacter sphaeroides.

Abstract

Genes coding for a classical membrane spanning chemoreceptor (mcpG) and a response regulator (cheY4) were identified in a region of Rhodobacter sphaeroides DNA unlinked to either of the two previously identified chemosensory operons. Immunogold electron microscopy had shown that the expression of chemoreceptors in R. sphaeroides varies with growth conditions. Using GFP fused to the newly identified McpG, we examined the targeting of this single methyl-accepting chemotaxis protein (MCP) under different growth conditions. The gene encoding the C-terminal McpG-GFP fusion was introduced by homologous recombination into the chromosome, replacing the wild-type gene. The resultant protein localized to the poles of the cell under aerobic, photoheterotrophic and anaerobic dark conditions, demonstrating that this MCP is expressed under all three growth conditions. More protein was always found at one pole than the other. The polar fluorescence increased during the cell cycle, with protein becoming evident at the second pole around the time of septation. At division, each daughter cell had a label at one pole, but the intensity of fluorescence was higher in the daughter cell containing the original labelled pole. McpG localization was not altered in a che Operon 1 deletion strain, lacking CheW1 and CheA1, but a che Operon 2 deletion strain, lacking CheW2, CheW3 and CheA2, showed significantly reduced polar localization. This observation indicates that polar localization of McpG depends on Che proteins encoded by Operon 2, but not homologues encoded by Operon 1.