Exploiting xylan as sugar donor for the synthesis of an antiproliferative xyloside using an enzyme cascade.

BACKGROUND: Currently, industrial societies are seeking for green alternatives to conventional chemical synthesis. This demand has merged with the efforts to convert lignocellulosic biomass into value-added products. In this context, xylan, as one of main components of lignocellulose, has emerged as a raw material with high potential for advancing towards a sustainable economy. RESULTS: In this study, the recombinant endoxylanase rXynM from the ascomycete Talaromyces amestolkiae has been heterologously expressed in Pichia pastoris and used as one of the catalysts of an enzyme cascade developed to synthesize the antiproliferative 2-(6-hydroxynaphthyl) ß-D-xylopyranoside, by transglycosylation of 2,6-dihydroxynaphthalene. The approach combines the use of two fungal xylanolytic enzymes, rXynM and the ß-xylosidase rBxTW1 from the same fungus, with the cost-effective substrate xylan. The reaction conditions for the cascade were optimized by a Central Composite Design. Maximal productions of 0.59 and 0.38 g/L were reached using beechwood xylan and birchwood xylan, respectively. For comparison, xylans from other sources were tested in the same reaction, suggesting that a specific optimization is required for each xylan variety. The results obtained using this enzyme cascade and xylan were similar or better to those previously reported for a single catalyst and xylobiose, an expensive sugar donor. CONCLUSIONS: Beechwood and birchwood xylan, two polysaccharides easily available from biomass, were used in a novel enzyme cascade to synthetize an antiproliferative agent. The approach represents a green alternative to the conventional chemical synthesis of 2-(6-hydroxynaphthyl) ß-D-xylopyranoside using a cost-effective substrate. The work highlights the role of xylan as a raw material for producing value-added products and the potential of fungal xylanolytic enzymes in the biomass conversion.

Bile-induced promoters for gene expression in Lactobacillus strains.

Bioengineering of probiotics allows the improvement of their beneficial characteristics. In this work, we develop a molecular tool that would allow the activation of desirable traits in probiotics once they reach the intestine. The activity of upstream regions of bile-inducible genes of Lactobacillus casei BL23 and Lactobacillus plantarum WCFS1 was analyzed using plasmids encoding an anaerobic fluorescent protein as reporter. The promoter P16090 from Lb. casei BL23 was selected and its bile induction confirmed in Lb. casei BL23, Lb. plantarum WCFS1, and in Lactobacillus rhamnosus and Lactobacillus reuteri strains. However, the induction did not occur in Lactococcus lactis MG1363 or Bifidobacterium strains. Studies with different bile compounds revealed the importance of cholic acid in the bile induction process. Induction of fluorescence was also confirmed for transformed Lb. casei BL23 under simulated colonic conditions and in the presence of intestinal microbiota. The developed vector, pNZ:16090-aFP, constitutes a promising tool suitable for the expression of genes of interest under intestinal conditions in probiotic strains of the species Lb. casei, Lb. plantarum, Lb. rhamnosus, and Lb. reuteri.