Identification of a Key Amino Acid of LuxS Involved in AI-2 Production in Campylobacter jejuni

Abstract

Autoinducer-2 (AI-2) mediated quorum sensing has been associated with the expression of virulence factors in a number of pathogenic organisms and has been demonstrated to play a role in motility and cytolethal distending toxin (cdt) production in Campylobacter jejuni. We have initiated the work to determine the molecular basis of AI-2 synthesis and the biological functions of quorum sensing in C. jejuni. In this work, two naturally occurring variants of C. jejuni 81116 were identified, one producing high-levels of AI-2 while the other is defective in AI-2 synthesis. Sequence analysis revealed a G92D mutation in the luxS gene of the defective variant. Complementation of the AI-2− variant with a plasmid encoded copy of the wild-type luxS gene or reversion of the G92D mutation by site-directed mutagenesis fully restored AI-2 production by the variant. These results indicate that the G92D mutation alone is responsible for the loss of AI-2 activity in C. jejuni. Kinetic analyses showed that the G92D LuxS has a ∼100-fold reduced catalytic activity relative to the wild-type enzyme. Findings from this study identify a previously undescribed amino acid that is essential for AI-2 production by LuxS and provide a unique isogenic pair of naturally occurring variants for us to dissect the functions of AI-2 mediated quorum sensing in Campylobacter.

Mining ofluxSgenes from rumen microbial consortia by metagenomic and metatranscriptomic approaches

“…In the present study, we demonstrated that S6, F7, H11, R39, N44 and C84 (amino acid numbering in LuxS of Vibrio harveyi) were conserved in LuxS homologues presumed form luxS gene sequences obtained from rumen microbial communities. Additionally, G86, which is a key amino acid in AI-2 production in bacteria (Plummer et al 2011), was also found in all ruminal LuxS homologues. Therefore, LuxS homologues presumed from full-length luxS gene sequences detected in this study would function as AI-2 synthase.…”

Critical Role of LuxS in the Virulence of Campylobacter jejuni in a Guinea Pig Model of Abortion

“…Natural transformation of the W7 and IA3902 strains with genomic DNA from a previous LuxS mutant developed in our laboratory was utilized to construct the W7⌬luxS and 3902⌬luxS strains, respectively, as previously described (38). For complementation of the luxS mutants, a copy of the luxS gene (including its putative promoter) was inserted into W7⌬luxS by homologous recombination into the intergenic region of the rRNA gene operon as described by Karlyshev and Wren with some modifications (30).…”

Polyphosphate-mediated modulation ofCampylobacter jejunibiofilm growth and stability

“…Planktonic cultures were tested after 22 h of growth at 42 °C with shaking, which was determined to be the optimal for AI-2 expression in C. jejuni in previous studies. 31 For static biofilm cultures: 3 mL of cultures were grown in borosilicate glass vials at 42 °C in duplicates and their supernatants pooled. Cultures were tested at 2 and 3 d of static biofilm growth.…”