Kinetics of H(2)O(2)-driven degradation of chitin by a bacterial lytic polysaccharide monooxygenase.

Lytic polysaccharide monooxygenases (LPMOs) catalyze the oxidative cleavage of glycosidic bonds in recalcitrant polysaccharides, such as cellulose and chitin, and are of interest in biotechnological utilization of these abundant biomaterials. It has recently been shown that LPMOs can use H(2)O(2), instead of O(2), as a cosubstrate. This peroxygenase-like reaction by a monocopper enzyme is unprecedented in nature and opens new avenues in chemistry and enzymology. Here, we provide the first detailed kinetic characterization of chitin degradation by the bacterial LPMO chitin-binding protein CBP21 using H(2)O(2) as cosubstrate. The use of (14)C-labeled chitin provided convenient and sensitive detection of the released soluble products, which enabled detailed kinetic measurements. The k(cat) for chitin oxidation found here (5.6 s(-1)) is more than an order of magnitude higher than previously reported (apparent) rate constants for reactions containing O(2) but no added H(2)O(2) The k(cat)/K(m) for H(2)O(2)-driven degradation of chitin was on the order of 10(6) m(-1) s(-1), indicating that LPMOs have catalytic efficiencies similar to those of peroxygenases. Of note, H(2)O(2) also inactivated CBP21, but the second-order rate constant for inactivation was about 3 orders of magnitude lower than that for catalysis. In light of the observed CBP21 inactivation at higher H(2)O(2) levels, we conclude that controlled generation of H(2)O(2)in situ seems most optimal for fueling LPMO-catalyzed oxidation of polysaccharides.