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Cloning, expression and characterization of two S-ribosylhomocysteine lyases from Lactobacillus plantarum YM-4-3: Implication of conserved and divergent roles in quorum sensing
- Source :
- Protein expression and purification. 145
- Publication Year :
- 2017
-
Abstract
- Quorum sensing (QS) is a means of cell-to-cell communication that regulates, via small signalling molecules, expression of a series of genes and controls multicellular behaviour in many bacterial species. The enzyme S-ribosylhomocysteine lyase (LuxS) transforms S-ribosylhomocysteine (SRH) into 4, 5-dihydroxy-2, 3-pentanedione (DPD), the precursor of the interspecies QS signalling molecule autoinducer-2 (AI-2). In this study, two LuxS-coding genes, luxS1 and luxS2, with 70% sequence identity were isolated from Lactobacillus plantarum YM-4-3, and overexpressed in Escherichia coli BL21 (DE3), and the protein products were purified successfully. After incubation of LuxS1 or LuxS2 with SRH, the reaction products were able to induce Vibrio harveyi BB170 bioluminescence, clearly demonstrating that both LuxS1 and LuxS2 synthesize AI-2 from SRH in vitro. Ellman's assay results revealed optimal temperatures for LuxS1 and LuxS2 of 45 and 37 °C, respectively, and their activities were stimulated or inhibited by several metal ions and chemical reagents. In addition, enzyme kinetics data showed that Km, Vmax and Kcat value of LuxS1 for the substrate (SRH) were higher than that of LuxS2. These results suggest that LuxS1 and LuxS2 mediate QS in a temperature-dependent manner and may play conserved roles in AI-2 synthesis but exhibit different activities in response to external environmental stress. To our knowledge, this paper is the first report of two luxS genes present in one bacterial genome and the subsequent comparative elucidation of their functions in AI-2 production. Collectively, our study provides a solid basis for future research concerning the AI-2/LuxS QS system in L. plantarum YM-4-3.
- Subjects :
- 0301 basic medicine
030106 microbiology
Gene Expression
medicine.disease_cause
Substrate Specificity
03 medical and health sciences
Lactones
Bacterial Proteins
medicine
Escherichia coli
Homoserine
Enzyme kinetics
Cloning, Molecular
Gene
Vibrio
chemistry.chemical_classification
biology
Vibrio harveyi
Temperature
Quorum Sensing
Gene Expression Regulation, Bacterial
biology.organism_classification
Lyase
Recombinant Proteins
Quorum sensing
Carbon-Sulfur Lyases
Enzyme
chemistry
Biochemistry
bacteria
Lactobacillus plantarum
Biotechnology
Subjects
Details
- ISSN :
- 10960279
- Volume :
- 145
- Database :
- OpenAIRE
- Journal :
- Protein expression and purification
- Accession number :
- edsair.doi.dedup.....00624bc98e30672bb6fd2d8c044e0aca