Application of xylanases and debranching enzymes of Cellulomonas sp. B6 for plant polysaccharides deconstruction into compounds with prebiotic and biopharmaceutical potential

ONTAÑON, O; GARRIDO, M; LANDONI M; TOPALIAN, JULIANA; NAVAS L; CAMPOS, E

Mendoza

Congreso; LVIII Annual Meeting of the Argentine Society for Biochemistry and Molecular Biology Research; 2022

SAIB

Xylan is the most abundant hemicellulose in nature, composed by a backbone of β 1-4 xylose unites and a diversity of substituent groups (arabinose, acetyl groups, glucuronic acids, feruloyl acids). The efficient degradation of xylan is of particular biotechnological interest since the hydrolysis products are desirable for a broad range of applications: short xylo-oligosaccharides (XOS) are emerging prebiotics, with demonstrated beneficial effects on health. Xylose, arabinose and glucuronic acids are precursors of biopharmaceuticals, food additives and biofuels. Cellulomonas sp. B6 is a soil bacterium with high xylanolytic potential, since it secretes a repertoire of xylanases by growth on biomass and encodes for diverse debranching enzymes active on hemicellulose. In this work we describe the catalytic potential of two xylanases from glycosode hydrolases (GH) families 10 and 11 (rCsXyn10A, rCsXyn11A), a GH62 α-L- arabinofuranosidase (rCsAbf62A) and a GH67 αglucuronidase (rCsAgu67A), expressed and purified as recombinant enzymes. Additionally, we studied the effect of combining these xylanases and debranching enzymes on biomass conversion to XOS and monomeric sugars. The hydrolysis products were analyzed by thin layer chromatography (TLC), highperformance anion-exchange chromatography/pulsed amperometric detection (HPAEC-PAD) and mass spectrometry (Maldi-TOF). The two recombinant xylanases showed different performance on commercial arabino- and glucuronoxylan, the main hemicelluloses present in plant biomass. The most active and thermostable was rCsXyn10A, which achieved a maximal activity above 400 U/mg on arabinoxylan at pH6 and 50°C, releasing short xylooligosaccharides (XOS) and xylose. Meanwhile, rCsXyn11A hydrolyzed xylans to xylosefree XOS reaching 65 U/mg in glucuronoxylan at 30°C, pH6. Debranching enzymes showed the expected activity: rCsAbf62A released arabinose from monosubstituted (α-1,2 and α-1,3) side-groups in arabinoxylooligosaccharides (AXOS) and arabinoxylan while rCsAgu67A was only active on external substitutions of glucuronic acid from glucuronoxylooligosaccharides (GXOS) but was not active on glucuronoxylan. Combined activity of each xylanase with rCsAbf62A improved the deconstruction of wheat arabinoxylan and wheat bran (a complex biomass composed of glucans, arabinoxylan, and lignin) to shorter XOS and xylobiose (X2). Besides, there was also a slight increase in the yield of xylose released by rCsXyn10A. rCsAgu67A was only active on oligosaccharides released by rCsXyn10A from glucuronoxylan-rich biomasses, obtaining glucuronic acid and unsubstituted XOS as products. Therefore, the individual or concerted activity of the tested enzymes has biotechnological potential for biomass hydrolysis into value added products. We will keep working in the combination of them and other enzymes in order to produce an enzyme cocktail with high activity and selectivity on hemicellulose.