Production of C1-oxidized cello-oligosaccharides by a fungal Lytic Polysaccharide Monooxygenase

GARRIDO, M; LANDONI, M.; GHIO, S; ONTAÑON, O; COUTO, A.S.; WIRTH, S; CAMPOS, E

Buenos Aires

Simposio; 3rd Argentinian Symposium on Glycobiology GlycoAR 2019; 2019

Lignocellulosic biomass has high potential as a renewableresource for the production of fuels and chemicals. However, itsnatural resistance to microbial and enzymatic deconstructionmakes its breakdown a slow and expensive process. A new classof secreted enzymes, known as lytic polysaccharidemonooxygenases (LPMOs) was recently identified due to itsboosting effect on enzymatic polysaccharide conversion byoxidizing glucose carbon atoms contained in cellulose, disruptingits crystalline structure and making it more accessible for glycosylhydrolases. Fungal LPMOs belong to family AA9 and can beclassified based on their regioselectivity (preferred site ofoxidation), as: type 1, which only oxidize C1; type 2, which onlyoxidize C4; and type 3, which can oxidize both C1 and C4. In thiswork, the coding sequence (960 bp) of an LPMO from the whiterot fungi Pycnoporus sanguienus, named AA9Ps3, was clonedwith its own signal sequence in fusion with a C-terminal 6-His tagin the expression vector pPIC9 and expressed in the yeast Pichiapastoris. The recombinant enzyme was purified from the culturebroth supernatant. Activity of AA9Ps3 on Phosphoric AcidSwollen Cellulose (PASC) resulted in the production of oxidizedcello-oligosaccharides, with degrees of polymerization 2 to 5,detected by HPAEC-PAD and MALDI-TOF MS. The m/z relationof the products obtained indicate the formation of aldonic acids,which correspond to the oxidation of glucose C1 atoms. AA9Ps3can be added to cellulolytic cocktails to improve cellulosebioconversion.