Data set for cloning and characterization of heterologous transporters in Saccharomyces cerevisiae and identification of important amino acids for xylose utilization

Abstract

The efficient uptake is important for the xylose utilization by Saccharomyces cerevisiae. A heterogenous transporter Mgt05196p was cloned from Meyerozyma guilliermondii and expressed in Saccharomyces cerevisiae[1]. This data article contains the transport characteristics of Mgt05196p in S. cerevisiae. The fluorescence of fusion protein Mgt05196p-GFP expressing strain was located on the cell surface demonstrated that the heterogenous transporter Mgt05196p was targeted to the plasma membrane of S. cerevisiae. The expressing of Mgt05196p in the hxt null S. cerevisiae endowed the strain with the glucose and d-xylose absorption capacity, as well as expressing the native d-xylose transporter Gal2p. The transmembrane domains of Mgt05196p were predicted and compared with the XylEp, whose crystal structure was revealed. And then, the homologous modeling of Mgt05196p was built basing on the XylEp to find out the crucial amino acid residues for sugars binding and transport.

Fig. 1

The fluorescence image of strain expressing fusion protein Mgt05196p-GFP and GFP. The Mgt05196p gene was fused with the GFP gene at the 3′ end and expressed in the hxt null strain EBY.VW4000. Unlike the GFP expressing strain, in which the fluorescence was dispersive, the fluorescence of the fusion protein Mgt05196p-GFP expressing strain was localized to the cell surface. Strains: the hxt null cells EBY.VW4000 expressed the GFP (reference) and GFP fusion protein Mgt05196p-GFP.

Fig. 2

The fermentation characteristics of the strains. The strain growth (A) and D-xylose consumption (B) in d-xylose aerobic fermentation. The strain growth (C), glucose consumption (D), d-xylose consumption (E), and ethanol production (F) in d-xylose and glucose oxygen-limited cofermentation. The 12 h precultured cells were collected, washed, and inoculated into 40 mL of SD medium plus 20 g L⁻¹ D-xylose or 20 g L⁻¹ D-xylose and 20 g L⁻¹ glucose, with an initial OD₆₀₀ of 20 (4.8 g DCW L⁻¹). The error bars represent the standard deviation of the biological triplicates. All of the strains were derived from the hxt null strain BSW4EYX, whose genome integrated the set of genes of XR, XDH, and XK. The control was strain BSW4PPX, which contained the empty plasmid for transporter expression. The other strains are represented by the transporters they expressed.

Fig. 3

The transmembrane domains prediction of transporters. (A), Protein: Mgt05196p. Length: 560; N-terminus: IN; Number of transmembrane helices: 12; Transmembrane helices: 52-76 107-124 135-152 165-182 193-210 227-244 327-345 354-371 382-400 421-445 456-474 487-506. (B) Protein: XylEp. Length: 491; N-terminus: IN; Number of transmembrane helices: 12; Transmembrane helices: 9-26 53-77 90-107 134-156 169-186 203-220 270-287 314-333 346-363 370-394 407-426 443-460.

Fig. 4

The homology 3 D model of Mgt05196p with D-xylose. The homology model of Mgt05196p was constructed according to the outward-facing and partly occluded structure of XylE using Discovery Studio. The location of D-xylose was determined using core-constrained protein docking and a modified CHARMm-based CDOCKER method. The best position among the 10 calculated positions of D-xylose was chosen by comparing the CDOCKER energies. The middle of the Mgt05196p model showed a colored D-xylose using the space-filling model. The residues of the predicted amino acids are also colorfully presented around D-xylose using the ball-and-stick model.

Fig. 5

The effect of mutating the amino acid residues of Mgt05196p to alanine (A) or some other residues (B) on the intracellular D-xylose accumulation. The well cultured cells were incubated in SD medium with 20 g L⁻¹D-xylose at 30 °C for 30, 60 and 120 min, then the intracellular D-xylose was extract using ddH₂O . The intracellular D-xylose was defined as the total amount of D-xylose and xylitol per gram in the dried cells. The error bars represent the standard deviation of the biological triplicates. All of the strains were derived from the hxt null strain EBY.VW4000. The control was strain BSW4PP, which contained the empty plasmid. The other strains are represented by the transporters and mutants they expressed.