Date of this Version
Lassalle, B. 2020. Investigation on the Structure-Function Relationships of Lysobacter antibioticus Phenazine O-methyltransferase - LaPhzM. Undergraduate Honors Thesis. University of Nebraska-Lincoln.
LaPhzM is an S-adenosyl-L-methionine (SAM) dependent, O-methyltransferase isolated from Lysobacter antibioticus, which catalyzes the biosynthesis of the phenazine antibiotic - myxin. The enzyme shares homology with O-methyltransferases of other soil microorganisms; however, the structure-function relationships of the enzyme have not yet been fully characterized. In this thesis work, site-directed mutagenesis was used to create six single amino acid substitutions (H251D, H251L, F151A, F138A, F138Y, and Y158F) based on the initial crystal structural analysis to test the functional roles of these residues in LaPhzM’s enzymatic activity. The results from this study confirmed that H251 is a residue for the catalysis of the enzyme. Our preliminary enzymatic assays suggested that F151, F138, and Y158 also contribute to mediating activity by non-covalent interactions with either the SAM cofactor or the phenazine substrate at the entry or within the binding pockets. Further characterization of the functional role of these residues at the active site of the enzyme is needed to elucidate the mechanism of action of LaPhzM. An in-depth mechanistic understanding of the enzyme may pave the way for utilizing this enzyme in producing derivatives of phenazine-based antibiotics.