Translocation and surface expression of lipidated serogroup B capsular Polysaccharide in Neisseria meningitidis.
The capsule of N. meningitidis serogroup B, (alpha2-->8)-linked polysialic acid and the capsules of other meningococcal serogroups and of other gram-negative bacterial pathogens are anchored in the outer membrane through a 1,2-diacylglycerol moiety. Previous work on the meningococcal cps complex in Escherichia coli K-12 indicated that deletion of genes designated lipA and lipB caused intracellular accumulation of hyperelongated capsule polymers lacking the phospholipid substitution. To better understand the role of lip and lipB in capsule expression in a meningococcal background, the location, sequence, and relationship to related bacterial capsule genes were defined and specific mutations in lipA and lipB were generated in the serogroup B meningococcal strain NMB. The lipA and lipB genes are located on the 3' end of the ctr operon and are most likely transcribed independently. Inactivation of lipA, lipB, and both resulted in the same total levels of capsular polymer production as in the parental controls; however, these mutants were as sensitive as an unencapsulated mutant to killing by normal human serum. Immunogold electron microscopy and flow cytometric analyses revealed intracellular inclusions of capsular polymers in lipA, lipB, and lipA lipB mutants. Capsular polymers purified from lipA, lipB, and lipA lipB mutants were lipidated. The phospholipid anchor was shown by gas chromatography-mass spectroscopy analysis to be a phosphodiester-linked 1,2-dipalmitoyl (C16:0) glycerol moiety and was identical in structure to that found on the wild-type meningococcal capsule polymers. Thus, lipA and lipB do not encode proteins responsible for diacylglycerophosphatidic acid substitution of the meningococcal capsule polymer; rather, they are required for proper translocation and surface expression of the lipidated polymer.
Tzeng
Yih-Ling
YL
Division of Infectious Diseases, Department of Medicine, Emory University School of Medicine, Atlanta, GA 30033, USA.
Datta
Anup K
AK
Strole
Cristy A
CA
Lobritz
Michael A
MA
Carlson
Russell W
RW
Stephens
David S
DS
D016667:Bacterial Capsules / Q000737:chemistry / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001770:Blood Bactericidal Activity; D005434:Flow Cytometry; D008401:Gas Chromatography-Mass Spectrometry; D006801:Humans; D007150:Immunohistochemistry; D008055:Lipids / Q000032:analysis; D009154:Mutation; D038541:Neisseria meningitidis, Serogroup B / Q000378:metabolism*; D010712:Phosphatidic Acids / Q000032:analysis; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D001426:Bacterial Proteins; D008055:Lipids; D010712:Phosphatidic Acids; C081999:lipA protein, Neisseria; C082000:lipB protein, Neisseria; C031599:dipalmitoylphosphatidic acid
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