UC Berkeley

ABSTRACT

Structure and Mechanism of Fungal Polysaccharide Monooxygenases

by

Elise Arielle Span

Doctor of Philosophy of Biophysics

University of California, Berkeley

Professor Michael A. Marletta, Chair

Polysaccharide monooxygenases (PMOs) are a newly discovered and growing superfamily of secreted copper catalysts found in nature. Initial PMO discoveries were made in 2011 describing the role of these enzymes in the cellulolytic machinery of fungi. Since then, additional families predominantly from fungi and bacteria have been characterized as having oxidative activity towards a variety of polysaccharide substrates. Chapter 1 introduces these enzymes and describes their origins, structure, function, as well as what can be surmised about their mechanism.

Chapter 2 describes a study that probes PMO mechanism in a system utilizing insoluble (and non-quantifiable) cellulose as a substrate. A combination of in vitro assays and electron paramagnetic resonance studies of a panel of single amino acid variants illustrates the function of the secondary coordination sphere in O2 activation by a cellulose-active PMO. This study bears directly on the recent controversy over the co-substrate of PMOs: although under some conditions PMOs can utilize peroxide, there is clearly a pathway for O2 utilization.

An example of discoveries that are on the horizon in the PMO field is described in Chapter 3. Genetic experiments and biochemical characterization of a homologously expressed PMO from Neurospora crassa shed light on a new family of PMOs that represents a significant departure from those studied previously. These enzymes are necessary for hyphal homing and fusion in filamentous fungi.