Biosynthesis of the Glucuronamide Moiety of the Capsular Polysaccharide of Campylobacter jejuni
Abstract
Campylobacter jejuni is a pathogenic organism that can cause campylobacteriosis in children and adults. Most commonly, Campylobacter infection is brought on by consumption of raw or undercooked poultry, unsanitary drinking water, or pet feces. Surrounding the C. jejuni bacterium is a coat of sugar molecules known as the capsular polysaccharide (CPS). The capsular polysaccharide can be very diverse among the different strains of C. jejuni, and this diversity is considered important for evading the host immune system. Modifications to the CPS of C. jejuni NCTC 11168 include O-methylation, phosphoramidylation, and amidation of glucuronate with either serinol or ethanolamine. The enzymes responsible for amidation of glucuronate are currently unknown. We functionally characterized five enzymes that are proposed to be required for glucuronamide biosynthesis in Campylobacter jejuni. Cj1441 was shown to catalyze the oxidation of UDP-α-D-glucose to UDP-α-D-glucuronic acid with NAD+ as the cofactor. The three-dimensional crystal structure of Cj1441 was determined in the presence of NAD+ and UDP-glucose bound in the active site of the enzyme. The amines required for amidation were synthesized from two PLP containing enzymes, Cj1436 and Cj1437. Decarboxylation of L-serine phosphate to ethanolamine phosphate is catalyzed by Cj1436. Cj1437 catalyzed the stereoselective transamination of dihydroxyacetone phosphate to (S)-serinol phosphate. Cj1438 was shown to catalyze amide bond formation using ATP, glucuronic acid, and either serinol phosphate or ethanolamine phosphate. Finally, Cj1435 catalyzed the hydrolysis of the phosphodiester of the glucuronamide of ethanolamine phosphate or the glucuronamide of (S)-serinol phosphate. This work represents the first known biosynthetic route for amide bond formation in capsular polysaccharides of C. jejuni.
Subject
Campylobacter jejuniBiochemistry
Enzymology
Capsular Polysaccharide
CPS
Kinetics
Proteins
Enzymes
Citation
Riegert, Alexander Sterling (2022). Biosynthesis of the Glucuronamide Moiety of the Capsular Polysaccharide of Campylobacter jejuni. Doctoral dissertation, Texas A&M University. Available electronically from https : / /hdl .handle .net /1969 .1 /198551.