Purification and Structural Characterization of the Auxiliary Activity 9 Native Lytic Polysaccharide Monooxygenase from Thermoascus aurantiacus and Identification of Its C1- and C4-Oxidized Reaction Products

Abstract

<p>Auxiliary activity 9 (AA9) lytic polysaccharide monooxygenases (LPMOs) are copperdependent<br>oxidoreductases that use O2 or H2O2 to perform oxidative cleavage of cellulose in the<br>presence of an electron donor. Combined with cellulases, they can assist in a more efficient cleavage<br>of cellulose. AA9 LPMOs have therefore attracted considerable attention in recent years for use in<br>biotechnological applications. Here, a native AA9 LPMO (nTaAA9A) from the thermophilic fungus<br>Thermoascus aurantiacus was purified and characterized. The enzyme was shown to be active and able<br>to cleave cellulose and xylan to produce C1- and C4-oxidized products. It was also found to retain<br>about 84.3, 63.7, and 35.3% of its activity after incubation for 30 min at 60, 70, and 80 C, respectively,<br>using quantitative activity determination. The structure was determined to 1.36 Å resolution and<br>compared with that of the recombinant enzyme expressed in Aspergillus oryzae. Structural differences<br>in the glycosylated Asn138 and in solvent-exposed loops were identified.<br></p>