Abstract
Pichia pastoris remains an amicable host for the expression of heterologous protein from eukaryotes. Achieving a high level of expression of yeast in the recombinant host has remained a challenge. Apart from the codon preference of genes and proper protein folding for efficient expression of the heterologous gene, the main bottleneck is to efficiently integrate the heterologous gene into the host DNA. The present study evaluated the choice of available transformation methods such as lithium chloride, electroporation, and spheroplast for efficient integration of xylose reductase (Xr) gene of Candida tropicalis in P. pastoris. The results revealed that electro transformation was found to be an efficient method for the transformation for xr in P. pastoris, yielding 1 x 106 transformants, followed by the lithium chloride method registering 2 x 103 transformants.
Highlights
D-Xylose is metabolized in yeast via., oxidoreductase pathway requiring several enzymes and cofactors
Xylose reductase (XR) is the first enzyme that converts D-xylose to xylitol requiring NAD(P)H, followed by the conversion of D-xylitol to D-xylulose by xylitol dehydrogenase (XDH) and requires NAD(P)+
Since xylitol production from yeast is less expensive than chemical methods, many studies focused on the biological production of xylitol
Summary
D-Xylose is metabolized in yeast via., oxidoreductase pathway requiring several enzymes and cofactors. Enzyme-based methods generate spheroplasts, which are plated in regeneration agar to obtain transformants. Lithium chloride mediated, electrophoration, and spheroplast methods were compared to transform P. pastoris GS115 with the Xr gene from Candida tropicalis cloned in vector pPIC3.5.
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