title: Total Biosynthesis of Legionaminic Acid, a Bacterial Sialic Acid Analogue.
abstract: Legionaminic acid, Leg5,7Ac2 , a nonulosonic acid like 5-acetamido neuraminic acid (Neu5Ac, sialic acid), is found in cell surface glycoconjugates of bacteria including the pathogens Campylobacter jejuni, Acinetobacter baumanii and Legionella pneumophila. The presence of Leg5,7Ac2 has been correlated with virulence in humans by mechanisms that likely involve subversion of the host's immune system or interactions with host cell surfaces due to its similarity to Neu5Ac. Investigation into its role in bacterial physiology and pathogenicity is limited as there are no effective sources of it. Herein, we construct a de novo Leg5,7Ac2 biosynthetic pathway by combining multiple metabolic modules from three different microbial sources (Saccharomyces cerevisiae, C. jejuni, and L. pneumophila). Over-expression of this de novo pathway in Escherichia coli that has been engineered to lack two native catabolic pathways, enables significant quantities of Leg5,7Ac2 (≈120 mg L(-1) of culture broth) to be produced. Pure Leg5,7Ac2 could be isolated and converted into CMP-activated sugar for biochemical applications and a phenyl thioglycoside for chemical synthesis applications. This first total biosynthesis provides an essential source of Leg5,7Ac2 enabling study of its role in prokaryotic and eukaryotic glycobiology.
authors.affiliation: Department of Chemistry and Biomolecular Sciences, Centre for Chemical and Synthetic Biology, University of Ottawa, Ottawa, ON, K1N 6N5, Canada.
authors.forename: Mohamed I
authors.initials: MI
authors.lastname: Hassan
authors.affiliation: Department of Chemistry and Biomolecular Sciences, Centre for Chemical and Synthetic Biology, University of Ottawa, Ottawa, ON, K1N 6N5, Canada.
authors.forename: Benjamin R
authors.initials: BR
authors.lastname: Lundgren
authors.affiliation: Department of Chemistry and Biomolecular Sciences, Centre for Chemical and Synthetic Biology, University of Ottawa, Ottawa, ON, K1N 6N5, Canada.
authors.forename: Michael
authors.initials: M
authors.lastname: Chaumun
authors.affiliation: Sussex Research Laboratories Inc., Ottawa, ON, K1A 0R6, Canada.
authors.forename: Dennis M
authors.initials: DM
authors.lastname: Whitfield
authors.affiliation: Sussex Research Laboratories Inc., Ottawa, ON, K1A 0R6, Canada.
authors.forename: Brady
authors.initials: B
authors.lastname: Clark
authors.affiliation: Human Health Therapeutics Portfolio, National Research Council Canada, Ottawa, ON, K1A 0R6, Canada. ian.schoenhofen@nrc-cnrc.gc.ca.
authors.forename: Ian C
authors.initials: IC
authors.lastname: Schoenhofen
authors.affiliation: Department of Chemistry and Biomolecular Sciences, Centre for Chemical and Synthetic Biology, University of Ottawa, Ottawa, ON, K1N 6N5, Canada. christopher.boddy@uottawa.ca.
authors.forename: Christopher N
authors.identifier: 0000-0002-3259-3184
authors.initials: CN
authors.lastname: Boddy
mesh_terms: D053898:Biosynthetic Pathways; D016123:Campylobacter jejuni / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D016952:Legionella pneumophila / Q000235:genetics / Q000378:metabolism*; D060847:Metabolic Engineering; D019158:N-Acetylneuraminic Acid / Q000031:analogs & derivatives*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D012794:Sialic Acids / Q000235:genetics / Q000378:metabolism*
chemical_list: C110570:5,7-diacetamido-8-O-acetyl-3,5,7,9-tetradeoxy-glycero-talo-nonulosonic acid; D012794:Sialic Acids; D019158:N-Acetylneuraminic Acid
keywords: glycobiology; glycoconjugates; legionaminic acid; synthetic biology; total biosynthesis