title: Specific amino acids of the glycosyltransferase LpsA direct the addition of glucose or galactose to the terminal inner core heptose of Haemophilus influenzae lipopolysaccharide via alternative linkages.
abstract: Lipopolysaccharide is the major glycolipid of the cell wall of the bacterium Haemophilus influenzae, a Gram-negative commensal and pathogen of humans. Lipopolysaccharide is both a virulence determinant and a target for host immune responses. Glycosyltransferases have high donor and acceptor substrate specificities that are generally limited to catalysis of one unique glycosidic linkage. The H. influenzae glycosyltransferase LpsA is responsible for the addition of a hexose to the distal heptose of the inner core of the lipopolysaccharide molecule and belongs to the glycosyltransferase family 25. The hexose added can be either glucose or galactose and linkage to the heptose can be either beta1-2 or beta1-3. Each H. influenzae strain uniquely produces only one of the four possible combinations of linked sugar in its lipopolysaccharide. We show that, in any given strain, a specific allelic variant of LpsA directs the anomeric linkage and the added hexose, glucose, or galactose. Site-directed mutagenesis of a single key amino acid at position 151 changed the hexose added in vivo from glucose to galactose or vice versa. By constructing chimeric lpsA gene sequences, it was shown that the 3' end of the gene directs the anomeric linkage (beta1-2 or beta1-3) of the added hexose. The lpsA gene is the first known example where interstrain variation in lipopolysaccharide core structure is directed by the specific sequence of a genetic locus encoding enzymes directing one of four alternative possible sugar additions from the inner core.
authors.affiliation: Department of Paediatrics, Weatherall Institute of Molecular Medicine, John Radcliffe Hospital, University of Oxford, Oxford OX3 9DS, United Kingdom. mary.deadman@paediatrics.ox.ac.uk
authors.forename: Mary E
authors.initials: ME
authors.lastname: Deadman
authors.forename: Susanna L
authors.initials: SL
authors.lastname: Lundström
authors.forename: Elke K H
authors.initials: EK
authors.lastname: Schweda
authors.forename: E Richard
authors.initials: ER
authors.lastname: Moxon
authors.forename: Derek W
authors.initials: DW
authors.lastname: Hood
mesh_terms: D000483:Alleles; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D002240:Carbohydrate Sequence; D020134:Catalytic Domain; D018387:Codon, Initiator; D005690:Galactose / Q000737:chemistry* / Q000378:metabolism; D014644:Genetic Variation; D005947:Glucose / Q000737:chemistry* / Q000378:metabolism; D006193:Haemophilus influenzae / Q000201:enzymology* / Q000235:genetics; D006539:Heptoses / Q000737:chemistry* / Q000378:metabolism; D006602:Hexosyltransferases / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D008070:Lipopolysaccharides / Q000096:biosynthesis* / Q000737:chemistry; D008969:Molecular Sequence Data; D013045:Species Specificity
chemical_list: D000596:Amino Acids; D001426:Bacterial Proteins; D018387:Codon, Initiator; D006539:Heptoses; D008070:Lipopolysaccharides; D006602:Hexosyltransferases; D005947:Glucose; D005690:Galactose