ABSTRACT
Pasteurella multocida (Pm) is a multi-species pathogen that causes diseases in animals and humans. Sialyltransferase activity has been detected in multiple Pm strains and sialylation has been shown to be important for the pathogenesis of Pm. Three putative sialyltransferase genes have been identified in Pm genomic strain Pm70. We have reported previously that a Pm0188 gene homolog in Pm strain P-1059 (ATCC 15742) encodes a multifunctional sialyltransferase (PmST1). We demonstrate here that while PmST1 prefers to use oligosaccharides as acceptors, PmST2 encoded by the Pm0508 gene homolog in the same Pm strain is a novel glycolipid α2-3-sialyltransferase that prefers to use lactosyl lipids as acceptor substrates. PmST2 and PmST1 thus complement each other for an efficient synthesis of α2-3-linked sialosides with or without lipid portion. In addition, ß1-4-linked galactosyl lipids are better PmST2 substrates than ß1-3-linked galactosyl lipids. PmST2 has been used successfully in the preparative scale synthesis of sialyllactosyl sphingosine (lyso-GM3), which is an important glycolipid and an intermediate for synthesizing more complex glycolipids such as gangliosides.
Subject(s)
Isoenzymes/metabolism , Pasteurella multocida , Recombinant Fusion Proteins/metabolism , Sialyltransferases/metabolism , Amino Acid Sequence , Animals , Chromatography, Affinity , Cloning, Molecular , Electrophoresis, Polyacrylamide Gel , Escherichia coli , G(M3) Ganglioside/analogs & derivatives , G(M3) Ganglioside/metabolism , Humans , Hydrogen-Ion Concentration , Isoenzymes/chemistry , Isoenzymes/genetics , Kinetics , Molecular Sequence Data , Oligosaccharides/metabolism , Pasteurella Infections/microbiology , Pasteurella multocida/enzymology , Pasteurella multocida/genetics , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/genetics , Sialyltransferases/chemistry , Sialyltransferases/genetics , Substrate Specificity , Transformation, BacterialABSTRACT
A series of α2-3-sialylated ß1-3-linked galactosides, including sialyl T-antigens, 3'-sialyl galacto-N-biose, 3'-sialyl lacto-N-biose, and their derivatives containing natural and non-natural sialic acid forms have been synthesized from simple monosaccharides using an efficient sequential two-step multienzyme approach.