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. 2002 May 1;363(Pt 3):437–447. doi: 10.1042/0264-6021:3630437

Characterization of a putative alpha-mannosyltransferase involved in phosphatidylinositol trimannoside biosynthesis in Mycobacterium tuberculosis.

Laurent Kremer 1, Sudagar S Gurcha 1, Pablo Bifani 1, Paul G Hitchen 1, Alain Baulard 1, Howard R Morris 1, Anne Dell 1, Patrick J Brennan 1, Gurdyal S Besra 1
PMCID: PMC1222496  PMID: 11964144

Abstract

Phosphatidyl-myo-inositol mannosides (PIMs), lipomannan (LM) and lipoarabinomannan (LAM) are an important class of bacterial factors termed modulins that are found in tuberculosis and leprosy. Although their structures are well established, little is known with respect to the molecular aspects of the biosynthetic machinery involved in the synthesis of these glycolipids. On the basis of sequence similarity to other glycosyltransferases and our previous studies defining an alpha-mannosyltransferase from Mycobacterium tuberculosis, named PimB [Schaeffer, Khoo, Besra, Chatterjee, Brennan, Belisle and Inamine (1999) J. Biol. Chem. 274, 31625-31631], which catalysed the formation of triacyl (Ac(3))-PIM(2) (i.e. the dimannoside), we have identified a related gene from M. tuberculosis CDC1551, now designated pimC. The use of a cell-free assay containing GDP-[(14)C]mannose, amphomycin and membranes from Myobacterium smegmatis overexpressing PimC led to the synthesis of a new alkali-labile PIM product. Fast-atom-bombardment MS established the identity of the new enzymically synthesized product as Ac(3)PIM(3) (i.e. the trimannoside). The results indicate that pimC encodes an alpha-mannosyltransferase involved in Ac(3)PIM(3) biosynthesis. However, inactivation of pimC in Myobacterium bovis Bacille Calmette-Guérin (BCG) did not affect the production of higher PIMs, LM and LAM when compared with wild-type M. bovis BCG, suggesting the existence of redundant gene(s) or an alternate pathway that may compensate for this PimC deficiency. Further analyses, which compared the distribution of pimC in a panel of M. tuberculosis strains, revealed that pimC was present in only 22% of the clinical isolates examined.

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Selected References

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  1. Abdian P. L., Lellouch A. C., Gautier C., Ielpi L., Geremia R. A. Identification of essential amino acids in the bacterial alpha -mannosyltransferase aceA. J Biol Chem. 2000 Dec 22;275(51):40568–40575. doi: 10.1074/jbc.M007496200. [DOI] [PubMed] [Google Scholar]
  2. Anthony L. S., Chatterjee D., Brennan P. J., Nano F. E. Lipoarabinomannan from Mycobacterium tuberculosis modulates the generation of reactive nitrogen intermediates by gamma interferon-activated macrophages. FEMS Immunol Med Microbiol. 1994 May;8(4):299–305. doi: 10.1111/j.1574-695X.1994.tb00456.x. [DOI] [PubMed] [Google Scholar]
  3. Apostolou I., Takahama Y., Belmant C., Kawano T., Huerre M., Marchal G., Cui J., Taniguchi M., Nakauchi H., Fournié J. J. Murine natural killer T(NKT) cells [correction of natural killer cells] contribute to the granulomatous reaction caused by mycobacterial cell walls. Proc Natl Acad Sci U S A. 1999 Apr 27;96(9):5141–5146. doi: 10.1073/pnas.96.9.5141. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Balasubramanian V., Pavelka M. S., Jr, Bardarov S. S., Martin J., Weisbrod T. R., McAdam R. A., Bloom B. R., Jacobs W. R., Jr Allelic exchange in Mycobacterium tuberculosis with long linear recombination substrates. J Bacteriol. 1996 Jan;178(1):273–279. doi: 10.1128/jb.178.1.273-279.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bateman Alex, Birney Ewan, Cerruti Lorenzo, Durbin Richard, Etwiller Laurence, Eddy Sean R., Griffiths-Jones Sam, Howe Kevin L., Marshall Mhairi, Sonnhammer Erik L. L. The Pfam protein families database. Nucleic Acids Res. 2002 Jan 1;30(1):276–280. doi: 10.1093/nar/30.1.276. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Belisle J. T., Vissa V. D., Sievert T., Takayama K., Brennan P. J., Besra G. S. Role of the major antigen of Mycobacterium tuberculosis in cell wall biogenesis. Science. 1997 May 30;276(5317):1420–1422. doi: 10.1126/science.276.5317.1420. [DOI] [PubMed] [Google Scholar]
  7. Besra G. S., Morehouse C. B., Rittner C. M., Waechter C. J., Brennan P. J. Biosynthesis of mycobacterial lipoarabinomannan. J Biol Chem. 1997 Jul 18;272(29):18460–18466. doi: 10.1074/jbc.272.29.18460. [DOI] [PubMed] [Google Scholar]
  8. Besra G. S. Preparation of cell-wall fractions from mycobacteria. Methods Mol Biol. 1998;101:91–107. doi: 10.1385/0-89603-471-2:91. [DOI] [PubMed] [Google Scholar]
  9. Bifani P. J., Mathema B., Liu Z., Moghazeh S. L., Shopsin B., Tempalski B., Driscol J., Frothingham R., Musser J. M., Alcabes P. Identification of a W variant outbreak of Mycobacterium tuberculosis via population-based molecular epidemiology. JAMA. 1999 Dec 22;282(24):2321–2327. doi: 10.1001/jama.282.24.2321. [DOI] [PubMed] [Google Scholar]
  10. Brennan P. J., Nikaido H. The envelope of mycobacteria. Annu Rev Biochem. 1995;64:29–63. doi: 10.1146/annurev.bi.64.070195.000333. [DOI] [PubMed] [Google Scholar]
  11. Brennan P., Ballou C. E. Biosynthesis of mannophosphoinositides by Mycobacterium phlei. The family of dimannophosphoinositides. J Biol Chem. 1967 Jul 10;242(13):3046–3056. [PubMed] [Google Scholar]
  12. Brosch R., Philipp W. J., Stavropoulos E., Colston M. J., Cole S. T., Gordon S. V. Genomic analysis reveals variation between Mycobacterium tuberculosis H37Rv and the attenuated M. tuberculosis H37Ra strain. Infect Immun. 1999 Nov;67(11):5768–5774. doi: 10.1128/iai.67.11.5768-5774.1999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Campbell J. A., Davies G. J., Bulone V., Henrissat B. A classification of nucleotide-diphospho-sugar glycosyltransferases based on amino acid sequence similarities. Biochem J. 1997 Sep 15;326(Pt 3):929–939. doi: 10.1042/bj3260929u. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Chatterjee D., Khoo K. H. Mycobacterial lipoarabinomannan: an extraordinary lipoheteroglycan with profound physiological effects. Glycobiology. 1998 Feb;8(2):113–120. doi: 10.1093/glycob/8.2.113. [DOI] [PubMed] [Google Scholar]
  15. Chatterjee D., Lowell K., Rivoire B., McNeil M. R., Brennan P. J. Lipoarabinomannan of Mycobacterium tuberculosis. Capping with mannosyl residues in some strains. J Biol Chem. 1992 Mar 25;267(9):6234–6239. [PubMed] [Google Scholar]
  16. Chatterjee D., Roberts A. D., Lowell K., Brennan P. J., Orme I. M. Structural basis of capacity of lipoarabinomannan to induce secretion of tumor necrosis factor. Infect Immun. 1992 Mar;60(3):1249–1253. doi: 10.1128/iai.60.3.1249-1253.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Cole S. T., Brosch R., Parkhill J., Garnier T., Churcher C., Harris D., Gordon S. V., Eiglmeier K., Gas S., Barry C. E., 3rd Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence. Nature. 1998 Jun 11;393(6685):537–544. doi: 10.1038/31159. [DOI] [PubMed] [Google Scholar]
  18. Dell A., Reason A. J., Khoo K. H., Panico M., McDowell R. A., Morris H. R. Mass spectrometry of carbohydrate-containing biopolymers. Methods Enzymol. 1994;230:108–132. doi: 10.1016/0076-6879(94)30010-0. [DOI] [PubMed] [Google Scholar]
  19. Ehlers M. R., Daffé M. Interactions between Mycobacterium tuberculosis and host cells: are mycobacterial sugars the key? Trends Microbiol. 1998 Aug;6(8):328–335. doi: 10.1016/s0966-842x(98)01301-8. [DOI] [PubMed] [Google Scholar]
  20. Flesselles B., Anand N. N., Remani J., Loosmore S. M., Klein M. H. Disruption of the mycobacterial cell entry gene of Mycobacterium bovis BCG results in a mutant that exhibits a reduced invasiveness for epithelial cells. FEMS Microbiol Lett. 1999 Aug 15;177(2):237–242. doi: 10.1111/j.1574-6968.1999.tb13738.x. [DOI] [PubMed] [Google Scholar]
  21. Geremia R. A., Petroni E. A., Ielpi L., Henrissat B. Towards a classification of glycosyltransferases based on amino acid sequence similarities: prokaryotic alpha-mannosyltransferases. Biochem J. 1996 Aug 15;318(Pt 1):133–138. doi: 10.1042/bj3180133. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Gilleron M., Ronet C., Mempel M., Monsarrat B., Gachelin G., Puzo G. Acylation state of the phosphatidylinositol mannosides from Mycobacterium bovis bacillus Calmette Guérin and ability to induce granuloma and recruit natural killer T cells. J Biol Chem. 2001 Jul 5;276(37):34896–34904. doi: 10.1074/jbc.M103908200. [DOI] [PubMed] [Google Scholar]
  23. Gordon S. V., Brosch R., Billault A., Garnier T., Eiglmeier K., Cole S. T. Identification of variable regions in the genomes of tubercle bacilli using bacterial artificial chromosome arrays. Mol Microbiol. 1999 May;32(3):643–655. doi: 10.1046/j.1365-2958.1999.01383.x. [DOI] [PubMed] [Google Scholar]
  24. Ha S., Walker D., Shi Y., Walker S. The 1.9 A crystal structure of Escherichia coli MurG, a membrane-associated glycosyltransferase involved in peptidoglycan biosynthesis. Protein Sci. 2000 Jun;9(6):1045–1052. doi: 10.1110/ps.9.6.1045. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Ho T. B., Robertson B. D., Taylor G. M., Shaw R. J., Young D. B. Comparison of Mycobacterium tuberculosis genomes reveals frequent deletions in a 20 kb variable region in clinical isolates. Yeast. 2000 Dec;17(4):272–282. doi: 10.1002/1097-0061(200012)17:4<272::AID-YEA48>3.0.CO;2-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Hoppe H. C., de Wet B. J., Cywes C., Daffé M., Ehlers M. R. Identification of phosphatidylinositol mannoside as a mycobacterial adhesin mediating both direct and opsonic binding to nonphagocytic mammalian cells. Infect Immun. 1997 Sep;65(9):3896–3905. doi: 10.1128/iai.65.9.3896-3905.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Hunter S. W., Brennan P. J. Evidence for the presence of a phosphatidylinositol anchor on the lipoarabinomannan and lipomannan of Mycobacterium tuberculosis. J Biol Chem. 1990 Jun 5;265(16):9272–9279. [PubMed] [Google Scholar]
  28. Jackson M., Raynaud C., Lanéelle M. A., Guilhot C., Laurent-Winter C., Ensergueix D., Gicquel B., Daffé M. Inactivation of the antigen 85C gene profoundly affects the mycolate content and alters the permeability of the Mycobacterium tuberculosis cell envelope. Mol Microbiol. 1999 Mar;31(5):1573–1587. doi: 10.1046/j.1365-2958.1999.01310.x. [DOI] [PubMed] [Google Scholar]
  29. Khoo K. H., Dell A., Morris H. R., Brennan P. J., Chatterjee D. Structural definition of acylated phosphatidylinositol mannosides from Mycobacterium tuberculosis: definition of a common anchor for lipomannan and lipoarabinomannan. Glycobiology. 1995 Feb;5(1):117–127. doi: 10.1093/glycob/5.1.117. [DOI] [PubMed] [Google Scholar]
  30. Kremer L., Baulard A., Estaquier J., Content J., Capron A., Locht C. Analysis of the Mycobacterium tuberculosis 85A antigen promoter region. J Bacteriol. 1995 Feb;177(3):642–653. doi: 10.1128/jb.177.3.642-653.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Mahenthiralingam E., Marklund B. I., Brooks L. A., Smith D. A., Bancroft G. J., Stokes R. W. Site-directed mutagenesis of the 19-kilodalton lipoprotein antigen reveals No essential role for the protein in the growth and virulence of Mycobacterium intracellulare. Infect Immun. 1998 Aug;66(8):3626–3634. doi: 10.1128/iai.66.8.3626-3634.1998. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Nigou J., Gilleron M., Cahuzac B., Bounéry J. D., Herold M., Thurnher M., Puzo G. The phosphatidyl-myo-inositol anchor of the lipoarabinomannans from Mycobacterium bovis bacillus Calmette Guérin. Heterogeneity, structure, and role in the regulation of cytokine secretion. J Biol Chem. 1997 Sep 12;272(37):23094–23103. doi: 10.1074/jbc.272.37.23094. [DOI] [PubMed] [Google Scholar]
  33. Pablos-Méndez A., Raviglione M. C., Laszlo A., Binkin N., Rieder H. L., Bustreo F., Cohn D. L., Lambregts-van Weezenbeek C. S., Kim S. J., Chaulet P. Global surveillance for antituberculosis-drug resistance, 1994-1997. World Health Organization-International Union against Tuberculosis and Lung Disease Working Group on Anti-Tuberculosis Drug Resistance Surveillance. N Engl J Med. 1998 Jun 4;338(23):1641–1649. doi: 10.1056/NEJM199806043382301. [DOI] [PubMed] [Google Scholar]
  34. Roach T. I., Barton C. H., Chatterjee D., Blackwell J. M. Macrophage activation: lipoarabinomannan from avirulent and virulent strains of Mycobacterium tuberculosis differentially induces the early genes c-fos, KC, JE, and tumor necrosis factor-alpha. J Immunol. 1993 Mar 1;150(5):1886–1896. [PubMed] [Google Scholar]
  35. Schaeffer M. L., Khoo K. H., Besra G. S., Chatterjee D., Brennan P. J., Belisle J. T., Inamine J. M. The pimB gene of Mycobacterium tuberculosis encodes a mannosyltransferase involved in lipoarabinomannan biosynthesis. J Biol Chem. 1999 Oct 29;274(44):31625–31631. doi: 10.1074/jbc.274.44.31625. [DOI] [PubMed] [Google Scholar]
  36. Schlesinger L. S., Hull S. R., Kaufman T. M. Binding of the terminal mannosyl units of lipoarabinomannan from a virulent strain of Mycobacterium tuberculosis to human macrophages. J Immunol. 1994 Apr 15;152(8):4070–4079. [PubMed] [Google Scholar]
  37. Shimada A., Ohta M., Iwasaki H., Ito E. The function of beta-N-acetyl-D-glucosaminyl monophosphorylundecaprenol in biosynthesis of lipoteichoic acids in a group of Bacillus strains. Eur J Biochem. 1988 Oct 1;176(3):559–565. doi: 10.1111/j.1432-1033.1988.tb14314.x. [DOI] [PubMed] [Google Scholar]
  38. Small P. M., McClenny N. B., Singh S. P., Schoolnik G. K., Tompkins L. S., Mickelsen P. A. Molecular strain typing of Mycobacterium tuberculosis to confirm cross-contamination in the mycobacteriology laboratory and modification of procedures to minimize occurrence of false-positive cultures. J Clin Microbiol. 1993 Jul;31(7):1677–1682. doi: 10.1128/jcm.31.7.1677-1682.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Snapper S. B., Melton R. E., Mustafa S., Kieser T., Jacobs W. R., Jr Isolation and characterization of efficient plasmid transformation mutants of Mycobacterium smegmatis. Mol Microbiol. 1990 Nov;4(11):1911–1919. doi: 10.1111/j.1365-2958.1990.tb02040.x. [DOI] [PubMed] [Google Scholar]
  40. Stover C. K., de la Cruz V. F., Fuerst T. R., Burlein J. E., Benson L. A., Bennett L. T., Bansal G. P., Young J. F., Lee M. H., Hatfull G. F. New use of BCG for recombinant vaccines. Nature. 1991 Jun 6;351(6326):456–460. doi: 10.1038/351456a0. [DOI] [PubMed] [Google Scholar]
  41. Venisse A., Berjeaud J. M., Chaurand P., Gilleron M., Puzo G. Structural features of lipoarabinomannan from Mycobacterium bovis BCG. Determination of molecular mass by laser desorption mass spectrometry. J Biol Chem. 1993 Jun 15;268(17):12401–12411. [PubMed] [Google Scholar]
  42. Videira P., Fialho A., Geremia R. A., Breton C., Sá-Correia I. Biochemical characterization of the beta-1,4-glucuronosyltransferase GelK in the gellan gum-producing strain Sphingomonas paucimobilis A.T.C.C. 31461. Biochem J. 2001 Sep 1;358(Pt 2):457–464. doi: 10.1042/0264-6021:3580457. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. van Embden J. D., Cave M. D., Crawford J. T., Dale J. W., Eisenach K. D., Gicquel B., Hermans P., Martin C., McAdam R., Shinnick T. M. Strain identification of Mycobacterium tuberculosis by DNA fingerprinting: recommendations for a standardized methodology. J Clin Microbiol. 1993 Feb;31(2):406–409. doi: 10.1128/jcm.31.2.406-409.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. van Soolingen D., de Haas P. E., Hermans P. W., Groenen P. M., van Embden J. D. Comparison of various repetitive DNA elements as genetic markers for strain differentiation and epidemiology of Mycobacterium tuberculosis. J Clin Microbiol. 1993 Aug;31(8):1987–1995. doi: 10.1128/jcm.31.8.1987-1995.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]

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