Cellulose-Binding Electron-Transfer Cytochrome Composing AA8 and CBM104 from the Wood-Rotting Fungus Armillaria tabescens.

Lytic polysaccharide monooxygenases (LPMOs) are activated by electrons supplied from external redox enzymes. Cellobiose dehydrogenase (CDH) and pyrroloquinoline quinone (PQQ)-dependent pyranose dehydrogenase (PDH) are electron donors to LPMOs. CDH and PDH each consist of two domains: a catalytic domain that oxidizes substrates to extract electrons, and an AA8 domain that receives and transfers these electrons to LPMOs. Although non-catalytic AA8 proteins comprising only an AA8 domain and a cellulose-binding module have been reported, their electron donors and physiological roles remain largely unclear. In this study, we characterized a protein from Armillaria tabescens (syn. Desarmillaria tabescens) that lacks a catalytic domain but includes both an AA8 domain and a cellulose-specific binding module, CBM104, which we designated AtAA8-CBM104. Electron-transfer assays demonstrated that the AA8 domain can accept electrons from an external AA12 catalytic domain, strongly suggesting that AtAA8-CBM104 act as an electron mediator transferring electrons from redox partners such as AA12 to adjacent LPMOs. Further, adsorption experiments showed that the CBM104 domain binds to crystalline cellulose but with lower adsorption efficiency than a previously characterized CBM104 appended to LPMO9. This observation suggests that despite AtAA8-CBM104 localization on cellulose being necessary for electron transfer to cellulose-bound LPMO9, excessive retention on the cellulose surface may interfere with LPMO9 catalysis. Furthermore, phylogenetic analysis indicated that AA8 appended to CBM104 lacking catalytic domains share similar electrostatic properties. These findings advance our understanding of non-catalytic AA8 proteins, underscoring their potential involvement in fungal redox networks linked to cellulose decomposition.