Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

LPMO AfAA9_B and Cellobiohydrolase AfCel6A from A. fumigatus Bost Enzymatic Saccharification Activity of Cellulase Cocktail

Version 1 : Received: 3 December 2020 / Approved: 6 December 2020 / Online: 6 December 2020 (18:15:08 CET)

A peer-reviewed article of this Preprint also exists.

Bernardi, A.V.; Gerolamo, L.E.; de Gouvêa, P.F.; Yonamine, D.K.; Pereira, L.M.S.; de Oliveira, A.H.C.; Uyemura, S.A.; Dinamarco, T.M. LPMO AfAA9_B and Cellobiohydrolase AfCel6A from A. fumigatus Boost Enzymatic Saccharification Activity of Cellulase Cocktail. Int. J. Mol. Sci. 2021, 22, 276. Bernardi, A.V.; Gerolamo, L.E.; de Gouvêa, P.F.; Yonamine, D.K.; Pereira, L.M.S.; de Oliveira, A.H.C.; Uyemura, S.A.; Dinamarco, T.M. LPMO AfAA9_B and Cellobiohydrolase AfCel6A from A. fumigatus Boost Enzymatic Saccharification Activity of Cellulase Cocktail. Int. J. Mol. Sci. 2021, 22, 276.

Journal reference: Int. J. Mol. Sci. 2020, 22, 276
DOI: 10.3390/ijms22010276

Abstract

Cellulose is the most abundant polysaccharide in lignocellulosic biomass, where it is interlinked with lignin and hemicellulose. Bioethanol can be produced from biomass. Because breaking down biomass is difficult, cellulose-active enzymes secreted by filamentous fungi play an important role in degrading recalcitrant lignocellulosic biomass. We characterized a cellobiohydrolase (AfCel6A) and lytic polysaccharide monooxygenase LPMO (AfAA9_B) from A. fumigatus after they were expressed in Pichia pastoris and purified. The biochemical parameters suggested that the enzymes were stable; the optimal temperature was ~60 °C. Further characterization revealed high turnover numbers (kcat of 147.9 s-1 and 0.64 s-1, respectively). Surprisingly, when combined, AfCel6A and AfAA9_B did not act synergistically. Association of AfCel6A and AfAA9_B inhibits the activity of AfCel6A, an outcome that needs to be further investigated. However, addition of AfCel6A or AfAA9_B boosts the enzymatic saccharification activity of a cellulase cocktail and the activity of cellulase Af-EGL7. The supplementation of an enzymatic cocktail with AfCel6A or AfAA9_B boosted the yield of fermentable sugars from complex substrates, especially sugarcane exploded bagasse, by up to 95%. The synergism between the cellulase cocktail and AfAA9_B is enzyme- and substrate-specific, which suggests a specific enzymatic cocktail for each biomass.

Subject Areas

GH6 cellobiohydrolase; AA9 LPMO; lignocellulose hydrolysis; bioethanol

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