A combined cell-consortium approach for lignocellulose degradation by specialized Lactobacillus plantarumcells

Abstract

BackgroundLactobacillus plantarum is an attractive candidate for metabolic engineering towards bioprocessing of lignocellulosic biomass to ethanol or polylactic acid, as its natural characteristics include high ethanol and acid tolerance and the ability to metabolize the two major polysaccharide constituents of lignocellulolytic biomass (pentoses and hexoses). We recently engineered L. plantarum via separate introduction of a potent cellulase and xylanase, thereby creating two different L. plantarum strains. We used these strains as a combined cell-consortium for synergistic degradation of cellulosic biomass.ResultsTo optimize enzymatic degradation, we applied the cell-consortium approach to assess the significance of enzyme localization by comparing three enzymatic paradigms prevalent in nature: (i) a secreted enzymes system, (ii) enzymes anchored to the bacterial cell surface and (iii) enzymes integrated into cellulosome complexes. The construction of the three paradigmatic systems involved the division of the production and organization of the enzymes and scaffold proteins into different strains of L. plantarum. The spatial differentiation of the components of the enzymatic systems alleviated the load on the cell machinery of the different bacterial strains. Active designer cellulosomes containing a xylanase and a cellulase were thus assembled on L. plantarum cells by co-culturing three distinct engineered strains of the bacterium: two helper strains for enzyme secretion and one producing only the anchored scaffoldin. Alternatively, the two enzymes were anchored separately to the cell wall. The secreted enzyme consortium appeared to have a slight advantage over the designer cellulosome system in degrading the hypochlorite pretreated wheat straw substrate, and both exhibited significantly higher levels of activity compared to the anchored enzyme consortium. However, the secreted enzymes appeared to be less stable than the enzymes integrated into designer cellulosomes, suggesting an advantage of the latter over longer time periods.ConclusionsBy developing the potential of L. plantarum to express lignocellulolytic enzymes and to control their functional combination and stoichiometry on the cell wall, this study provides a step forward towards optimal biomass bioprocessing and soluble fermentable sugar production. Future expansion of the preferred secreted-enzyme and designer-cellulosome systems to include additional types of enzymes will promote enhanced deconstruction of cellulosic feedstocks.Electronic supplementary materialThe online version of this article (doi:10.1186/1754-6834-7-112) contains supplementary material, which is available to authorized users.

Toward combined delignification and saccharification of wheat straw by a laccase-containing designer cellulosome

“…Interestingly, members of the family-6 glycoside hydrolase enzymes have never been found to be part of any known cellulosome system. However, endocellulase Tfu_1074 (Cel6A) was demonstrated to confer enhanced cellulose-degrading activity in simple designer cellulosomes (38,53). In contrast, Tfu_0620 (Cel6B) is an exoglucanase, which was shown to cause an antiproximity effect in designer cellulosomes, and was therefore deemed incompatible with the cellulosomal mode (38).…”

Display of recombinant proteins at the surface of lactic acid bacteria: strategies and applications

“…In a very recent study, Morais et al used the versatility of the pSIP expression system to produce what could be called the first functional minicellulosome in LAB [139]. In one part of this study, strains producing cellwall anchored variants of the previously used xylanase and cellulase were generated using previously developed LPxTG anchors [41].…”

Lactic Acid Production from Lignocellulose – A Review of Major Challenges and Selected Solutions

“…Engineered L. plantarum with Cel8A endoglucanase obtained from C. thermocellum was able to grow on cellooligosaccharides up to 5–6 glucose residues long . Simple consortia of recombinant L. plantarum that secreted cellulase‐xylanase mixtures to breakdown wheat straw to fermentable sugar have been created . A L. lactis strain that secreted endoglucanase and β ‐glucosidase was developed through the construction of an artificial operon.…”