Characterization of a GH5 endoxylanase from Penicillium funiculosum and its synergism with GH16 endo-1,3(4)-glucanase in saccharification of sugarcane bagasse

Abstract

Abstract The production of second-generation fuels from lignocellulosic residues such as sugarcane bagasse (SCB) requires the synergistic interaction of key cellulose-degrading enzymes and accessory proteins for their complete deconstruction to useful monomeric sugars. Here, we recombinantly expressed and characterized unknown GH5 xylanase from P. funiculosum ( Pf Xyn5) in Pichia pastoris , which was earlier found in our study to be highly implicated in SCB saccharification. The Pf Xyn5 has a molecular mass of ~ 55 kDa and showed broad activity against a range of substrates like xylan, xyloglucan, laminarin and p-nitrophenyl-β- d -xylopyranoside, with the highest specific activity of 0.7 U/mg against xylan at pH 4.5 and 50 °C. Analysis of the degradation products of xylan and SCB by Pf Xyn5 showed significant production of xylooligosaccharides (XOS) with a degree of polymerization (DP) ranging from two (DP 2 ) to six (DP 6 ), thus, suggesting that the Pf Xyn5 is an endo-acting enzyme. The enzyme synergistically improved the saccharification of SCB when combined with the crude cellulase cocktail of P. funiculosum with a degree of synergism up to 1.32. The Pf Xyn5 was further expressed individually and simultaneously with a notable GH16 endoglucanase ( Pf Egl16) in a catabolite-derepressed strain of P. funiculosum , Pf Mig1 88 , and the saccharification efficiency of the secretomes from the resulting transformants were investigated on SCB. The secretome of Pf Mig1 88 overexpressing Xyn5 or Egl16 increased the saccharification of SCB by 9% or 7%, respectively, over the secretome of Pf Mig1 88 , while the secretome of dual transformant increased SCB saccharification by ~ 15% at the same minimal protein concentration.