Quantifying Oxidation of Cellulose-Associated Glucuronoxylan by Two Lytic Polysaccharide Monooxygenases from Neurospora crassa
| Autor: | Lisbeth Olsson, Dejan M. Petrović, Heidi Østby, Anikó Várnai, Olav A. Hegnar, Vincent G. H. Eijsink |
|---|---|
| Rok vydání: | 2021 |
| Předmět: |
lytic polysaccharide monooxygenases
Polysaccharide Applied Microbiology and Biotechnology xylan Mixed Function Oxygenases Neurospora crassa Fungal Proteins Cell wall chemistry.chemical_compound lignocellulose Glucuronoxylan Hemicellulose LPMO Enzymology and Protein Engineering Cellulose Phylogeny chemistry.chemical_classification Ecology biology hemicellulose biology.organism_classification Xylan glucuronoxylan chemistry Biochemistry Xylans Oxidation-Reduction Function (biology) Food Science Biotechnology |
| Zdroj: | Applied and Environmental Microbiology |
| ISSN: | 1098-5336 0099-2240 |
| DOI: | 10.1128/aem.01652-21 |
| Popis: | Family AA9 lytic polysaccharide monooxygenases (LPMOs) are abundant in fungi, where they catalyze oxidative depolymerization of recalcitrant plant biomass. These AA9 LPMOs cleave cellulose and some also act on hemicelluloses, primarily other (substituted) β-(1→4)-glucans. Oxidative cleavage of xylan has been shown for only a few AA9 LPMOs, and it remains unclear whether this activity is a minor side reaction or primary function. Here, we show that Neurospora crassa LPMO9F (NcLPMO9F) and the phylogenetically related, hitherto uncharacterized NcLPMO9L from N. crassa are active on both cellulose and cellulose-associated glucuronoxylan but not on glucuronoxylan alone. A newly developed method for simultaneous quantification of xylan-derived and cellulose-derived oxidized products showed that NcLPMO9F preferentially cleaves xylan when acting on a cellulose–beechwood glucuronoxylan mixture, yielding about three times more xylan-derived than cellulose-derived oxidized products. Interestingly, under similar conditions, NcLPMO9L and the previously characterized McLPMO9H, from Malbranchea cinnamomea, showed different xylan-to-cellulose preferences, giving oxidized product ratios of about 0.5:1 and 1:1, respectively, indicative of functional variation among xylan-active LPMOs. Phylogenetic and structural analysis of xylan-active AA9 LPMOs led to the identification of characteristic structural features, including unique features that do not occur in phylogenetically remote AA9 LPMOs, such as four AA9 LPMOs whose lack of activity toward glucuronoxylan was demonstrated in the present study. Taken together, the results provide a path toward discovery of additional xylan-active LPMOs and show that the huge family of AA9 LPMOs has members that preferentially act on xylan. These findings shed new light on the biological role and industrial potential of these fascinating enzymes. IMPORTANCE Plant cell wall polysaccharides are highly resilient to depolymerization by hydrolytic enzymes, partly due to cellulose chains being tightly packed in microfibrils that are covered by hemicelluloses. Lytic polysaccharide monooxygenases (LPMOs) seem well suited to attack these resilient copolymeric structures, but the occurrence and importance of hemicellulolytic activity among LPMOs remain unclear. Here, we show that certain AA9 LPMOs preferentially cleave xylan when acting on a cellulose–glucuronoxylan mixture, and that this ability is the result of protein evolution that has resulted in a clade of AA9 LPMOs with specific structural features. Our findings strengthen the notion that the vast arsenal of AA9 LPMOs in certain fungal species provides functional versatility and that AA9 LPMOs may have evolved to promote oxidative depolymerization of a wide variety of recalcitrant, copolymeric plant polysaccharide structures. These findings have implications for understanding the biological roles and industrial potential of LPMOs. |
| Databáze: | OpenAIRE |
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