Pplement four). Notably, N. crassa expresses a putative intracellular -xylosidase, GH43-2 (NCU01900), when grown on xylan (Sun et al., 2012). Purified GH43-2 displayed robust hydrolase activity towards xylodextrins using a degree of polymerization (DP) spanning from two to 8, and with a pH optimum close to 7 (Figure MEK Inhibitor Molecular Weight 1–figure supplement 5). The outcomes with CDT-2 and GH43-2 confirm these obtained independently in Cai et al. (2014). As with cdt-1, orthologues of cdt-2 are widely distributed inside the fungal kingdom (Galazka et al., 2010), suggesting that lots of fungi consume xylodextrins derived from plant cell walls. Moreover, as with intracellular -glucosidases (Galazka et al., 2010), intracellular -xylosidases are also widespread in fungi (Sun et al., 2012) (Figure 1–figure supplement six). Cellodextrins and xylodextrins derived from plant cell walls are not catabolized by wild-type S. cerevisiae (Matsushika et al., 2009; Galazka et al., 2010; Young et al., 2010). Reconstitution of a cellodextrin transport and consumption pathway from N. crassa in S. cerevisiae enabled this yeast to ferment cellobiose (Galazka et al., 2010). We consequently reasoned that expression of a functional xylodextrin transport and consumption system from N. crassa could possibly further expand the capabilities ofLi et al. eLife 2015;four:e05896. DOI: ten.7554/eLife.2 ofResearch articleComputational and systems biology | EcologyFigure 1. Consumption of xylodextrins by engineered S. cerevisiae. (A) Two oligosaccharide components derived in the plant cell wall. Cellodextrins, derived from cellulose, are a major source of glucose. Xylodextrins, derived from hemicellulose, are a major supply of xylose. The 6-methoxy group (blue) distinguishes glucose derivatives from xylose. R1, R2 = H, cellobiose or xylobiose; R1 = -1,4-linked glucose monomers in cellodextrins of bigger degrees of polymerization; R2 = -1,4-linked xylose monomers in xylodextrins of mAChR5 Agonist drug larger degrees of polymerization. (B) Xylose and xylodextrins remaining within a culture of S. cerevisiae grown on xylose and xylodextrins and expressing an XR/XDH xylose consumption pathway, CDT-2, and GH43-2, with a beginning cell density of OD600 = 1 below aerobic conditions. (C) Xylose and xylodextrins inside a culture as in (B) but with a starting cell density of OD600 = 20. In both panels, the concentrations of xylose (X1) and xylodextrins with greater DPs (X2 five) remaining inside the culture broth following distinct periods of time are shown. All experiments had been performed in biological triplicate, with error bars representing standard deviations. DOI: ten.7554/eLife.05896.003 The following figure supplements are offered for figure 1: Figure supplement 1. Transcriptional levels of transporters expressed in N. crassa grown on diverse carbon sources. DOI: 10.7554/eLife.05896.004 Figure supplement 2. Development of N. crassa strains on unique carbon sources. DOI: ten.7554/eLife.05896.005 Figure supplement 3. Xylodextrins in the xylan culture supernatant of the N. crassa cdt-2 strain. DOI: 10.7554/eLife.05896.006 Figure supplement 4. Transport of xylodextrins in to the cytoplasm of S. cerevisiae strains expressing N. crassa transporters. DOI: 10.7554/eLife.05896.007 Figure supplement 5. Xylobiase activity on the predicted -xylosidase GH43-2. DOI: 10.7554/eLife.05896.008 Figure supplement 6. Phylogenetic distribution of predicted intracellular -xylosidases GH43-2 in filamentous fungi. DOI: 10.7554/eLife.05896.009 Figure supplement 7. Xylodextrin consumption profi.
