Abstract
Bacteria have evolved a wide range of chemoreceptors with different ligand specificities. Typically, chemoreceptors bind ligands with elevated specificity and ligands serve as growth substrates. However, there is a chemoreceptor family that has a broad ligand specificity including many compounds that are not of metabolic value. To advance the understanding of this family, we have used the PcaY_PP (PP2643) chemoreceptor of Pseudomonas putida KT2440 as a model. Using Isothermal Titration Calorimetry we showed here that the recombinant ligand binding domain (LBD) of PcaY_PP recognizes 17 different C6-ring containing carboxylic acids with KD values between 3.7 and 138 μM and chemoeffector affinity correlated with the magnitude of the chemotactic response. Mutation of the pcaY_PP gene abolished chemotaxis to these compounds; phenotype that was restored following gene complementation. Growth experiments using PcaY_PP ligands as sole C-sources revealed functional relationships between their metabolic potential and affinity for the chemoreceptor. Thus, only 7 PcaY_PP ligands supported growth and their KD values correlated with the length of the bacterial lag phase. Furthermore, PcaY_PP ligands that did not support growth had significantly higher KD values than those that did. The receptor has thus binds preferentially compounds that serve as C-sources and amongst them those that rapidly promote growth. Tightest binding compounds were quinate, shikimate, 3-dehydroshikimate and protocatechuate, which are at the interception of the biosynthetic shikimate and catabolic quinate pathways. Analytical ultracentrifugation studies showed that ligand free PcaY_PP-LBD is present in a monomer-dimer equilibrium (KD = 57.5 μM). Ligand binding caused a complete shift to the dimeric state, which appears to be a general feature of four-helix bundle LBDs. This study indicates that the metabolic potential of compounds is an important parameter in the molecular recognition by broad ligand range chemoreceptors.
Highlights
Bacteria need to constantly adapt to changing environmental conditions to assure survival
Previous research has shown that most individual chemoreceptor ligand binding domain (LBD) fold into stable domains that bind their chemoeffectors with the same affinity as the full-length receptor (Milligan and Koshland, 1993; Matilla and Krell, 2017)
In the model organism of this study, P. putida KT2440, eight chemoreceptors have been functionally characterized (Supplementary Table 1). These receptors were shown to be activated by the direct binding of 34 different chemoeffectors, which are all of metabolic value since they support growth as Cand/or N-sources
Summary
Bacteria need to constantly adapt to changing environmental conditions to assure survival These adaptations are achieved through a variety of signal transduction systems that most commonly include one- and two-component systems as well as chemosensory pathways (Galperin, 2005; Laub and Goulian, 2007; Hazelbauer et al, 2008). Escherichia coli is the traditional model to study chemosensory signaling mechanisms. This bacterium has four chemoreceptors and an aerotaxis receptor that feed into a single chemotaxis pathway (Parkinson et al, 2015). Several alternative model organisms have been established to investigate chemotaxis in bacteria with lifestyles different to that of the enterobacterium E. coli
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