ABSTRACT
Resolving the enzymatic pathways leading to sialic acids (Sias) in bacteria are vitally important for understanding their roles in pathogenesis and for subsequent development of tools to combat infections. A detailed characterization of the involved enzymes is also essential due to the highly applicable properties of Sias, i.e., as used in a wide range of medical applications and human nutrition. Bacterial strains that produce Sias display them mainly on their cell surface to mimic animal cells thereby evading the host's immune system. Despite several studies, little is known about the virulence mechanisms of the fish pathogen Aliivibrio salmonicida. The genome of A. salmonicida LFI1238 contains a gene cluster homologous to the Escherichia coli neuraminic acid (Neu) gene cluster involved in biosynthesis of Sias found in the E. coli capsule. This cluster is probably responsible for the biosynthesis of Neu found in A. salmonicida. In this work, we have produced and characterized the sialic acid (Sia) synthase NeuB1, the key enzyme in the pathway. The Sia synthase is an enzyme producing N-acetylneuraminic acid by the condensation of N-acetylmannosamine and phosphoenolpyruvate. Genome content, kinetic data obtained, together with structural considerations, have led us to the prediction that the substrate for NeuB1 from A. salmonicida, E. coli and Streptococcus agalactiae among others, is 4-O-acetyl-N-acetylmannosamine. This means that the product of its enzymatic reaction is 7-O-acetyl-N-acetylneuraminic acid. We propose a pathway for production of this Sia in A. salmonicida, and present evidence for the presence of diacetylated Neu in the bacterium.
