UC Berkeley

Polysaccharide Monooxygenases (PMOs) activate molecular oxygen through a copper active site to hydroxylate and subsequently cleave polysaccharides. These enzymes are of interest for their potential role in biofuel production and C-H activation chemistry. The biological role of many of the first described members of this superfamily family was in polysaccharide breakdown for carbon uptake carried out by fungi and bacteria. More recently, roles for these enzymes outside of carbon utilization have emerged. This dissertation will focus on two fungal PMOs and one bacterial PMO that fulfill roles other than catabolism. Chapter 1 outlines the current understanding of PMOs as a superfamily of enzymes. This introduction details the copper active site, known substrates of PMOs, and what is known about the steps involved in catalysis. The following three chapters focus on putative PMOs that have unique biological roles other than nutrient degradation for uptake by the organism. Chapter 2 focuses on the biochemical characterization of the PMO, CWR-1, and how it plays a role in the allorecognition process of Neurospora crassa. Evidence shows that CWR-1 enzymatic activity is not crucial for this role but is likely a PPI. Chapter 3 focuses on the putative PMO, HAM-7. This PMO is involved in fungal development, and N. crassa strains that lack this protein have severe growth phenotypes. The chapter details efforts to express and purify this protein to obtain more detailed biochemical information. Chapter 4 focuses on the Vibrio cholerae PMO, GBPA, and the proteases VcLAP and VcLAPX. GBPA has been shown to play a role in adherence to mucin during the infection lifecycle of the bacteria Vibrio cholerae. The potential activity of GBPA on mucin and the characterization of these proteases is presented.