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In addition to utilizing glycosylated phosphatidylinositols (GPIs) as anchors for surface proteins, protozoan parasites of the genus Leishmania synthesize two novel classes of GPI: the polydisperse lipophosphoglycans (LPGs) and a family of low molecular weight glycoinositol phospholipids (GIPLs). We now show that LPG is expressed in high copy number (6 x 10(6) molecules/cell) in the promastigote (insect) stage of L. donovani but not in the amastigote stage, which infects mammalian macrophages. Detection of these molecules was by gas chromatography-mass spectrometric analyses and by a sensitive radiolabeling procedure. In contrast, a novel family of GIPLs was present in high copy number (approximately 10(7) molecules/cell) in both promastigote and amastigote stages of L. donovani. These glycolipids were purified and analyzed by gas chromatography-mass spectrometry, methylation analysis, and by chemical and enzymatic sequencing after deamination and NaB3H4 reduction. Promastigotes contained three major GIPLs species with the following generalized structure [formula: see text] where R = H for isoM2, Man α1- for isoM3 or Man α1-2Man α1- for isoM4. Amastigotes contained two major GIPL species that lacked the α1-3-linked mannose branch and had the linear structures Man α1-6Man α1-4GlcN (M2) and Man α1-2Man α1-6Man α1-4GlcN (M3) linked to alkylacyl-PI. The 1-O-alkyl-2-acyl-PI moieties of all these species contained predominantly C18:0 alkyl chains and C16:0 or C18:0 fatty acids. Amastigotes contained, in addition, a GalNAc β1-3 terminating glycosphingolipid with homology to the mammalian para Forssman glycolipid. This glycolipid appeared to be a constituent of the parasite membrane but was not metabolically labeled with [3H]glucose, suggesting that it was acquired from host cells. These results suggest that LPG may not be required for amastigote survival in the mammalian host and that the GIPLs are likely to be major components on the surface membrane in both stages.

NCBI PubMed ID: 1831200
Journal NLM ID: 2985121R
Publisher: Baltimore, MD: American Society for Biochemistry and Molecular Biology
Institutions: Department of Biochemistry, University of Dundee, United Kingdom

Journal NLM ID: 2985121R
Publisher: Baltimore, MD: American Society for Biochemistry and Molecular Biology
Methods: 1H NMR, FAB-MS, GC-MS

PapG is the adhesin at the tip of the P pilus that mediates attachment of uropathogenic Escherichia coli to the uroepithelium of the human kidney. The human specific allele of PapG binds to globoside (GbO4), which consists of the tetrasaccharide GalNAc β 1-3Gal α 1-4Gal β 1-4Glc linked to ceramide. Here, we present the crystal structures of a binary complex of the PapG receptor binding domain bound to GbO4 as well as the unbound form of the adhesin. The biological importance of each of the residues involved in binding was investigated by site-directed mutagenesis. These studies provide a molecular snapshot of a host-pathogen interaction that determines the tropism of uropathogenic E. coli for the human kidney and is critical to the pathogenesis of pyelonephritis.

chemistry, Bacterial, metabolism, microbiology, pathogenicity, molecular, X-ray, Escherichia coli, Female, Molecular Sequence Data, proteins, mutagenesis, protein structure, models, amino acid sequence, Sequence Alignment, kidney, Fimbriae, Protein Binding, Crystallography, Humans, Adhesins, Binding Sites, Escherichia coli Infections, Crystallization, Fimbriae Proteins, Globosides, Protein Conformation, Tertiary, Pyelonephritis, Urothelium

NCBI PubMed ID: 11440716
Journal NLM ID: 0413066
Publisher: Cambridge, MA: Cell Press
Institutions: Department of Molecular Microbiology, Washington University School of Medicine, Saint Louis, MO 63110, USA
Methods: MAD