title: Molecular characterization and subcellular localization of macrophage infectivity potentiator, a Chlamydia trachomatis lipoprotein.
abstract: Macrophage infectivity potentiator (MIP) was originally reported to be a chlamydial lipoprotein from experiments showing incorporation of radiolabeled palmitic acid into native and recombinant MIP; inhibition of posttranslational processing of recombinant MIP by globomycin, known to inhibit signal peptidase II; and solubility of native MIP in Triton X-114. However, the detailed structural characterization of the lipid moiety on MIP has never been fully elucidated. In this study, bioinformatics and mass spectrometry analysis, as well as radiolabeling and immunochemical experiments, were conducted to further characterize MIP structure and subcellular localization. In silico analysis showed that the amino acid sequence of MIP is conserved across chlamydial species. A potential signal sequence with a contained lipobox was identified, and a recombinant C20A variant was prepared by replacing the probable lipobox cysteine with an alanine. Both incorporation of U-(14)C-esterified glycerol and [U-(14)C]palmitic acid and posttranslational processing that was inhibitable by globomycin were observed for recombinant wild-type MIP but not for the recombinant C20A MIP variant. The fatty acid contents of native and recombinant MIP were analyzed by gas chromatography-mass spectrometry, and the presence of amide-linked fatty acids in recombinant MIP was investigated by alkaline methanolysis. These results demonstrated a lipid modification in MIP similar to that of other prokaryotic lipoproteins. In addition, MIP was detected in an outer membrane preparation of Chlamydia trachomatis elementary bodies and was shown to be present at the surfaces of elementary bodies by surface biotinylation and surface immunoprecipitation experiments.
authors: Division of Rheumatology, Department of Internal Medicine, University Hospital, 1211 Geneva 14, Switzerland.
authors: Laurence
authors: L
authors: Neff
authors: Sawsan
authors: S
authors: Daher
authors: Patrick
authors: P
authors: Muzzin
authors: Ursula
authors: U
authors: Spenato
authors: Fazil
authors: F
authors: Gülaçar
authors: Cem
authors: C
authors: Gabay
authors: Sylvette
authors: S
authors: Bas
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002692:Chlamydia trachomatis / Q000235:genetics / Q000378:metabolism*; D019295:Computational Biology; D008401:Gas Chromatography-Mass Spectrometry; D015151:Immunoblotting; D047468:Immunoprecipitation; D008074:Lipoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D019308:Palmitic Acid / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid
chemical_list: D001426:Bacterial Proteins; D008074:Lipoproteins; D011993:Recombinant Fusion Proteins; D019308:Palmitic Acid