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Biochemical Characterization of the Putative Glucosyltransferase, WapB, Which Acts on the Uncapped Core Oligosaccharide of Pseudomonas aeruginosa

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Title: Biochemical Characterization of the Putative Glucosyltransferase, WapB, Which Acts on the Uncapped Core Oligosaccharide of Pseudomonas aeruginosa
Author: El-Osta, Lana
Department: Department of Molecular and Cellular Biology
Program: Molecular and Cellular Biology
Advisor: Kimber, Matthew
Abstract: Pseudomonas aeruginosa is a Gram-negative, opportunistic pathogen known to cause disease in Cystic Fibrosis patients. Lipopolysaccharide (LPS) forms the outermost layer of the outer membrane and is an important virulence factor. LPS is divided conceptually into three domains: lipid A, core oligosaccharide, and (sometimes) O-antigen. Uncapped (O-antigen free) core is often modified with a terminal glucose (GlcIV) on L-RhaA; while a wapB mutant produces LPS lacking GlcIV. At present, little is known regarding the enzymatic properties of WapB. My hypothesis is that WapB is a β-1,2-glucosyltransferase that transfers GlcIV to the uncapped core OS of P. aeruginosa PAO1. WapB proved to be insoluble when expressed, and several solubilization strategies were explored. Of these strategies, membrane extraction using the detergent DDM proved the most promising, suggesting WapB interacts tightly with the membrane. Complementation was also used to show that minimum 20 C-terminal residues and a DXD motif are required for function.
URI: http://hdl.handle.net/10214/14205
Date: 2018-08-27


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