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Τετάρτη 9 Αυγούστου 2017

Expression, Docking, and Molecular Dynamics of Endo-β-1,4-xylanase I Gene of Trichoderma virens in Pichia stipitis

It is essential that major carbohydrate polymers in the lignocellulosic biomass are converted into fermentable sugars for the economical production of energy. Xylan, the major component of hemicelluloses, is the second most naturally abundant carbohydrate polymer comprising 20–40% of the total biomass. Endoxylanase (EXN) hydrolyzes xylan into mixtures of xylooligosaccharides. The objective of this study was to genetically modify Pichia stipitis, a pentose sugar fermenting yeast species, to hydrolyze xylan into xylooligosaccharides via cloning and heterologous extracellular expression of EXNI gene from locally isolated Trichoderma virens species. Pichia stipitis was engineered to carry the EXNI gene of T. virens using pGAPZα expression vector. The open reading frame encodes 191 amino acids and SDS-PAGE analysis revealed a 24 kDA recombinant protein. The EXNI activity expressed by recombinant P. stipitis clone under standard conditions using 1% beechwood xylan was 31.7 U/ml. Molecular docking and molecular dynamics simulations were performed to investigate EXNI-xylan interactions. Free EXNI and xylan bound EXNI exhibited similar stabilities and structural behavior in aqueous medium. Furthermore, this in silico work opens avenues for the development of newer generation EXN proteins that can perform better and have enhanced catalytic activity.

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