THE IMPACT OF GLYCOSYLATION ON ACTIVE SITE STRUCTURE AND ACTIVITY IN THE FUNGUS ENZYME MOLOX

Abstract

Lipoxygenases from pathogenic fungi belong to the lipoxygenase family that catalyze the C-H activation of polyunsaturated fatty acids to form diverse hydroperoxides. While the lipoxygenase catalytic domains are structurally and functionally similar, the fungal enzymes are decorated with N-linked glycosylations. MoLOX, a lipoxygenase from the fungus M. oryzae, is emerging as an important target for the devastating rice blast disease. Here we demonstrate for the first time that hydrogen transfer, associated with C-H cleavage of linoleic acid by MoLOX, occurs by a hydrogen tunneling mechanism. Using the temperature dependent kinetic isotope effect, [delta]Ea, as a kinetic reporter of tunneling efficiency, the loss of N-linked carbohydrates is linked to an increase in the activation energy for deuterium transfer, consistent with an impairment of the tunneling ready state. These results have important implications for MoLOX inhibitor design towards a potential 'treatment' of rice blast disease.