New insights into hydroxyectoine synthesis and its transcriptional regulation in the broad‐salt growing halophilic bacteriumChromohalobacter salexigens

Abstract

Elucidating the mechanisms controlling the synthesis of hydroxyectoine is important to design novel genetic engineering strategies for optimizing the production of this biotechnologically relevant compatible solute. The genome of the halophilic bacterium Chromohalobacter salexigens carries two ectoine hydroxylase genes, namely ectD and ectE, whose encoded proteins share the characteristic consensus motif of ectoine hydroxylases but showed only a 51.9% identity between them. In this work, we have shown that ectE encodes a secondary functional ectoine hydroxylase and that the hydroxyectoine synthesis mediated by this enzyme contributes to C. salexigens thermoprotection. The evolutionary pattern of EctD and EctE and related proteins suggests that they may have arisen from duplication of an ancestral gene preceding the directional divergence that gave origin to the orders Oceanospirillales and Alteromonadales. Osmoregulated expression of ectD at exponential phase, as well as the thermoregulated expression of ectD at the stationary phase, seemed to be dependent on the general stress factor RpoS. In contrast, expression of ectE was always RpoS-dependent regardless of the growth phase and osmotic or heat stress conditions tested. The data presented here suggest that the AraC-GlxA-like EctZ transcriptional regulator, whose encoding gene lies upstream of ectD, plays a dual function under exponential growth as both a transcriptional activator of osmoregulated ectD expression and a repressor of ectE transcription, privileging the synthesis of the main ectoine hydroxylase EctD. Inactivation of ectZ resulted in a higher amount of the total ectoines pool at the expenses of a higher accumulation of ectoine, with maintenance of the hydroxyectoine levels. In addition to the transcriptional control, our results suggest a strong post-transcriptional regulation of hydroxyectoine synthesis. Data on the accumulation of ectoine and hydroxyectoine in rpoS and ectZ strains pave the way for using these genetic backgrounds for metabolic engineering for hydroxyectoine production.

Draft Genomes of Halophilic Chromohalobacter and Halomonas Strains Isolated from Brines of The Carpathian Foreland, Poland

“…However, both Halomonas strains, 11-S5 and 25-S5, had a higher number of genes belonging to this category (99 and 101 genes, respectively) than isolates belonging to the genus Chromohalobacter , which consisted of 69-89 such genes, depending on the strain. This observation is consistent with previous reports that Halomonas and Chromohalobacter have a high diversity of biosynthetic processes 10 , 12 , 13 Based on these results, the annotation of BGCs with antiSMASH was performed. Interestingly, as a result, more BGCs were identified in Chromohalobacter strains than Halomonas , despite a smaller number of genes associated with processes identified during the analysis.…”

Guilty by association: Importers, exporters and MscS‐type mechanosensitive channels encoded in biosynthetic gene clusters for the stress‐protectant ectoine

“…Ectoine biosynthetic genes are typically organized in an evolutionarily conserved and osmotically inducible operon ( ectABC ), which might also comprise the gene for the ectoine hydroxylase (EctD) (Argandona et al, 2021; Bursy et al, 2007), and/or that for a specialized aspartokinase (Ask_Ect), an enzyme enhancing the synthesis of the ectoine biosynthetic precursor l ‐aspartate‐β‐semialdehyde (Hermann et al, 2020; Stöveken et al, 2011). These gene clusters are also often associated with a gene encoding a MarR‐type repressor (EctR) controlling ect gene transcription (Mustakhimov et al, 2010).…”

“…Synthesis of hydroxyectoine by the ectoine hydroxylase EctD is dependent on the prior production of ectoine (Hermann et al, 2020; Höppner et al, 2014). Microorganisms that possess EctD can either produce a mixture of the two ectoines or synthesize exclusively hydroxyectoine (Argandona et al, 2021; Czech, Hermann, et al, 2018; Pastor et al, 2010). We found that H. neptunium and Novosphingobium sp.…”

Microbial conversion of carbon dioxide and hydrogen into the fine chemicals hydroxyectoine and ectoine

“…Salinity is a requirement to promote the expression of ect genes (Argandoña et al, 2021). None of the strains tested could grow at salinities above 8% NaCl.…”