Evidence for Pentapeptide-Dependent and Independent CheB Methylesterases

Abstract

Many bacteria possess multiple chemosensory pathways that are composed of homologous signaling proteins. These pathways appear to be functionally insulated from each other, but little information is available on the corresponding molecular basis. We report here a novel mechanism that contributes to pathway insulation. We show that, of the four CheB paralogs of Pseudomonas aeruginosa PAO1, only CheB2 recognizes a pentapeptide at the C-terminal extension of the McpB (Aer2) chemoreceptor (KD = 93 µM). McpB is the sole chemoreceptor that stimulates the Che2 pathway, and CheB2 is the methylesterase of this pathway. Pectobacterium atrosepticum SCRI1043 has a single CheB, CheB_Pec, and 19 of its 36 chemoreceptors contain a C-terminal pentapeptide. The deletion of cheB_Pec abolished chemotaxis, but, surprisingly, none of the pentapeptides bound to CheB_Pec. To determine the corresponding structural basis, we solved the 3D structure of CheB_Pec. Its structure aligned well with that of the pentapeptide-dependent enzyme from Salmonella enterica. However, no electron density was observed in the CheB_Pec region corresponding to the pentapeptide-binding site in the Escherichia coli CheB. We hypothesize that this structural disorder is associated with the failure to bind pentapeptides. Combined data show that CheB methylesterases can be divided into pentapeptide-dependent and independent enzymes.

Chemotaxis of the Human Pathogen Pseudomonas aeruginosa to the Neurotransmitter Acetylcholine

“…S6 ). Of the 36 chemoreceptors of SCRI1043, two (ECA_RS10935 and ECA_RS05475) possess a dCache type LBD ( 61 ). The LBDs of these receptors share only 20% and 17% sequence identity with PctD, respectively ( Fig.…”

A Selective Tether Recruits Activated Response Regulator CheB to Its Chemoreceptor Substrate

“…Linkers in response regulators are conformationally flexible. They can be without regular secondary structure or be partly or entirely helical in different crystallographic or functional forms ( 16 , 28 , 29 ). The CheB linker has little regular secondary structure in the closed form, but secondary structure prediction ( 30 ) suggested that helix α5 could extend past its closed conformation termination at residue 132 to include residues through 138 in the linker, thus incorporating what had been nonhelical residues 133 and 134 plus the single helical turn from residues 135 to 138.…”

“…Related carboxyl-terminal pentapeptides are found on ∼11% chemoreceptor sequences (>25,000 at the time of the analysis), distributed among many bacterial species and 11 bacterial phyla ( 15 ). These carboxyl-terminal sequences presumably bind to cognate CheBs, an interaction that has been documented in the only other species investigated besides E. coli ( 16 ).…”

“…These interactions are incompletely defined. The low-activity, i.e., unphosphorylated, form of CheB from E. coli or Pseudomonas aeruginosa binds to the pentapeptide with weak dissociation constants of 130 to 160 μM ( 21 ) and ∼90 μM ( 16 ), respectively, implying that the pentapeptide could not serve as an effective tether to increase the local concentration of the enzyme. Binding of high-activity, phosphorylated CheB to the pentapeptide has not been measured.…”

Pectobacterium brasiliense 1692 Chemotactic Responses and the Role of Methyl-Accepting Chemotactic Proteins in Ecological Fitness

“…Furthermore, unlike Dickeya, chemotaxis is less studied for most Pectobacterium spp. (Antúnez-Lamas et al, 2009a,b;Río-Álvarez et al, 2015;Velando et al, 2020). Toward this end, our first aim was to determine Pb 1692 responses to various ligands.…”