Date Available

4-22-2021

Year of Publication

2021

Degree Name

Doctor of Philosophy (PhD)

Document Type

Doctoral Dissertation

College

Pharmacy

Department/School/Program

Pharmaceutical Sciences

First Advisor

Dr. Steven G. Van Lanen

Abstract

Antibiotic-resistance has become a widespread problem in the United States and across the globe. Meanwhile, new antibiotics are entering the clinic at an alarmingly low rate. Highly-modified nucleosides, a class of natural products often produced by actinobacteria, target MraY bacterial translocase I. MraY is a clinically unexploited enzyme target that is ubiquitous and essential to peptidoglycan cell wall biosynthesis. The nucleoside antibiotics known vary in efficacy and the functionalities contributing to improved activity is poorly understood. Sphaerimicin, a newly discovered modified nucleoside, has potent inhibitory activity with an IC50 of 13.65 nM against MraY. In general, sphaerimicin is primarily effective against gram-positive bacteria (MIC ranges from 2-16 μg/mL against Enterococcus faecium and Staphylococcus aureus), but little is known about the biosynthesis and mechanism of action. Sphaerimicin has highly unusual structural features, including a heavily modified ribosylated glycl-uridine disaccharide core that is appended to a dihydroxylated piperidine ring.

The novel biosynthesis of these features was investigated, leading to the functional characterization of six enzymes critical for the disaccharide core formation from uridine monophosphate. Recently, a unique S-adenosylmethionine- and PLP-dependent alpha-aminobutyric acid transferase and a nonheme Fe(II)- and alpha-ketogluterate-dependent hydroxylase from the sphaerimicin biosynthetic pathway were characterized. This part of the pathway extends the carbon scaffold, which will be crucial for formation of the piperidine ring. Not only does this study provide new chemical entities to help better understand MraY as a target, it could potentially reveal new enzymatic chemistries that will power innovative chemoenzymatic synthesis and genome mining to uncover new natural products.

Digital Object Identifier (DOI)

https://doi.org/10.13023/etd.2021.151

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