Structural Insights into Inhibition of Bacillus anthracis Sporulation by a Novel Class of Non-heme Globin Sensor Domains

Abstract

Pathogenesis by Bacillus anthracis requires coordination between two distinct activities: plasmid-encoded virulence factor expression (which protects vegetative cells from immune surveillance during outgrowth and replication) and chromosomally encoded sporulation (required only during the final stages of infection). Sporulation is regulated by at least five sensor histidine kinases that are activated in response to various environmental cues. One of these kinases, BA2291, harbors a sensor domain that has ϳ35% sequence identity with two plasmid proteins, pXO1-118 and pXO2-61. Because overexpression of pXO2-61 (or pXO1-118) inhibits sporulation of B. anthracis in a BA2291-dependent manner, and pXO2-61 expression is strongly up-regulated by the major virulence gene regulator, AtxA, it was suggested that their function is to titrate out an environmental signal that would otherwise promote untimely sporulation. To explore this hypothesis, we determined crystal structures of both plasmid-encoded proteins. We found that they adopt a dimeric globin fold but, most unusually, do not bind heme. Instead, they house a hydrophobic tunnel and hydrophilic chamber that are occupied by fatty acid, which engages a conserved arginine and chloride ion via its carboxyl head group. In vivo, these domains may therefore recognize changes in fatty acid synthesis, chloride ion concentration, and/or pH. Structure-based comparisons with BA2291 suggest that it binds ligand and dimerizes in an analogous fashion, consistent with the titration hypothesis. Analysis of newly sequenced bacterial genomes points to the existence of a much broader family of non-heme, globin-based sensor domains, with related but distinct functionalities, that may have evolved from an ancestral heme-linked globin.

A Dual Role for the Bacillus anthracis Master Virulence Regulator AtxA: Control of Sporulation and Anthrax Toxin Production

“…Our transcriptional profiling studies reported previously ( Bourgogne et al, 2003 ) revealed that a pXO2 gene, pXO2-0075 , is positively regulated 54-fold by atxA . The predicted amino acid sequence of pX02-0075 exhibits high sequence similarity to the signal sensor domain of the BA2291 sporulation sensor histidine kinase, which is a key component of the sporulation phosphorelay ( White et al, 2006 ; Scaramozzino et al, 2009 ; Stranzl et al, 2011 ). White et al (2006) showed that over-expression of pXO2-0075 in a Sterne-like strain resulted in a marked decrease in sporulation that was suppressed by deletion of the sensor histidine kinase BA2291.…”

Two Surfaces of a Conserved Interdomain Linker Differentially Affect Output from the RST Sensing Module of the Bacillus subtilis Stressosome

“…However, the question of whether the nonheme globin domain of RsbRA is a sensor of acute environmental stress is unresolved. Characterization of two similar nonheme globin domains that modulate sporulation in Bacillus anthracis suggests that they are capable of sensing fatty acids and chloride via binding within a distinctive tunnel and chamber (41). The lack of these structural features seems to preclude the B. subtilis domain from sensing similar signals, and the cellular parameter to which it responds remains unknown.…”

Heme-based Globin-coupled Oxygen Sensors: Linking Oxygen Binding to Functional Regulation of Diguanylate Cyclase, Histidine Kinase, and Methyl-accepting Chemotaxis

“…Globin sensor domains that lack heme binding are known, although their functions have not been determined (45,46). However, given the accumulating information about the O 2 -mediated catalytic regulation and physicochemical properties of GCSs, it is worth summarizing specific characteristics of GCSs in comparison with those of non-GCS heme-based oxygen sensors.…”