A point mutation in the active site of Legionella pneumophila O-acetyltransferase results in modified lipopolysaccharide but does not influence virulence.
The majority of clinical isolates of Legionella pneumophila serogroup 1 produce lipopolysaccharide (LPS) that reacts with monoclonal antibody (MAb) 3/1. By using a negative cell sorting method, we isolated a spontaneous LPS mutant from L. pneumophila serogroup 1 strain Corby that lost reactivity with this MAb. The mutant contained a single nucleotide exchange in position 169 of the lag-1 gene that encodes an O-acetyltransferase that is responsible for O-acetylation of the L. pneumophila O-repeat unit (legionaminic acid). This mutation resulted in a single amino acid exchange in a highly conserved motif present in many O-acetyltransferase-like proteins. RT-PCR analysis revealed that the mutant lag-1 gene was transcribed, but the resulting protein lacked O-acetyltransferase activity. Chemical analysis of the mutant LPS revealed that it lacked 8-O-acetyl groups in legionaminic acid. In addition, the mutant failed to produce high-molecular-weight long-chain O-polysaccharide. Complementation of the mutant with the wild-type lag-1 gene restored reactivity with MAb 3/1 and the chemical structure of the wild-type LPS. Strain Corby and its MAb 3/1-negative mutant were indistinguishable in their serum resistance characteristics, and in uptake and intracellular multiplication in Acanthamoeba castellanii and macrophages.
Lück
P C
PC
Institut für Medizinische Mikrobiologie und Hygiene, TU Dresden, Germany. Christian.Lueck@mailbox.tu-dresden.de
Freier
T
T
Steudel
C
C
Knirel
Y A
YA
Lüneberg
E
E
Zähringer
U
U
Helbig
J H
JH
D000048:Acanthamoeba / Q000382:microbiology; D000123:Acetyltransferases / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D000939:Epitopes; D005816:Genetic Complementation Test; D006168:Guinea Pigs; D006801:Humans; D016952:Legionella pneumophila / Q000235:genetics* / Q000276:immunology / Q000472:pathogenicity; D008070:Lipopolysaccharides / Q000276:immunology / Q000378:metabolism*; D016676:Macrophages, Alveolar / Q000382:microbiology; D017354:Point Mutation; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D012794:Sialic Acids / Q000276:immunology / Q000378:metabolism; D020298:U937 Cells; D014774:Virulence / Q000235:genetics
C110570:5,7-diacetamido-8-O-acetyl-3,5,7,9-tetradeoxy-glycero-talo-nonulosonic acid; D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; D000939:Epitopes; D008070:Lipopolysaccharides; D012794:Sialic Acids; D000123:Acetyltransferases