Biosynthesis of Mycobacterial Lipoarabinomannan

Besra, Gurdyal S.; Morehouse, Caroline B.; Rittner, Christian M.; Waechter, Charles J.; Brennan, Patrick J.

doi:10.1074/jbc.272.29.18460

Cited by 160 publications

(191 citation statements)

References 49 publications

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“…These results, indicating that LMs contain the same PI unit forms as ManLAMs in the same ratio, further support a biosynthetic link between LMs and ManLAMs, recently suggested by Besra et al [23].…”

Section: Discussionsupporting

confidence: 89%

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Lipoarabinomannans: characterization of the multiacylated forms of the phosphatidyl-myo-inositol anchor by NMR spectroscopy

Nigou

Gilleron

Puzo

1999

Biochemical Journal

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Lipoarabinomannans, which exhibit a large spectrum of immunological activities, emerge as the major antigens of mycobacterial envelopes. The lipoarabinomannan structure is based on a phosphatidyl-myo-inositol anchor whose integrity has been shown to be crucial for lipoarabinomannan biological activity and particularly for presentation to CD4/CD8 double-negative αβT cells by CD1 molecules. In this report, an analytical approach was developed for high-resolution 31P-NMR analysis of native, i.e. multiacylated, lipoarabinomannans. The one-dimensional 31P spectrum of cellular lipoarabinomannans, from Mycobacterium bovis Bacillus Calmette–Guérin, exhibited four 31P resonances typifying four types of lipoarabinomannans. Two-dimensional 1H-31P heteronuclear multiple-quantum-correlation/homonuclear Hartmann–Hahn analysis of the native molecules showed that these four types of lipoarabinomannan differed in the number and localization of fatty acids (from 1 to 4) esterifying the anchor. Besides the three acylation sites previously described, i.e. positions 1 and 2 of glycerol and 6 of the mannosyl unit linked to the C-2 of myo-inositol, we demonstrate the existence of a fourth acylation position at the C-3 of myo-inositol. We report here the first structural study of native multiacylated lipoarabinomannans, establishing the structure of the intact phosphatidyl-myo-inositol anchor. Our findings would help gain more understanding of the molecular basis of lipoarabinomannan discrimination in the binding process to CD1 molecules.

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Section: Discussionsupporting

confidence: 89%

“…the well-described phosphatidyl-myo-inositol mannosides (PIMs) [22][23][24]. The multiacylated nature of PIMs and LMs has been recently re-examined by others [17,22] and us (M. Gilleron, J. Nigou, B. Cahuzac and G. Puzo, unpublished work) in an attempt to find out, by structure homologies, whether they are biosynthetic precursors of LAMs.…”

Section: Introductionmentioning

confidence: 99%

Lipoarabinomannans: characterization of the multiacylated forms of the phosphatidyl-myo-inositol anchor by NMR spectroscopy

Nigou

Gilleron

Puzo

1999

Biochemical Journal

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“…On the basis of the literature precedent for the use of photoprobes incorporating benzophenone, a unique series of compounds were designed specifically to probe the biosynthesis of the mannan portion of LM and LAM, through the utilization of PPM synthase and a PPM-dependent α(1 → 6)mannosyltransferase. The probes are chemically novel in that they incorporate a flexible linker unit attached to the photophore, reducing the isoprene-related structure from 7 to 10 units in mycobacteria [17] to just 2 units thereby removing the need for elaborate syntheses of modified isoprenoid units. The results were very gratifying in relation to substrate specificity and photoinhibition.…”

Section: Discussionmentioning

confidence: 99%

“…Intuitively, it should proceed via the sequential addition of Manp residues to PI to produce PIMs (2-6 Manp residues) and LM (up to 20 Manp residues), which is then extensively arabinosylated to form AraLAM. The biosynthetic relationship of PI → PIMs → LM → AraLAM has recently been supported by biochemical [17] and genetic studies [18][19][20][21]. Besra et al [17] demonstrated that Ac 1 PIM 2 (according to the nomenclature of Kordulakova et al [21]) is specifically extended by an addition of Manp residues from the alkali-stable sugar donor C 50 /C 40 /C 35 -PPM (where PPM stands for polyprenol monophosphomannose) to form higher PIMs and further to a 'linear' α(1 → 6)-LM via a PPM-dependent α(1 → 6)mannosyltransferase.…”

Section: Introductionmentioning

confidence: 99%

“…The biosynthetic relationship of PI → PIMs → LM → AraLAM has recently been supported by biochemical [17] and genetic studies [18][19][20][21]. Besra et al [17] demonstrated that Ac 1 PIM 2 (according to the nomenclature of Kordulakova et al [21]) is specifically extended by an addition of Manp residues from the alkali-stable sugar donor C 50 /C 40 /C 35 -PPM (where PPM stands for polyprenol monophosphomannose) to form higher PIMs and further to a 'linear' α(1 → 6)-LM via a PPM-dependent α(1 → 6)mannosyltransferase. The generation of PPM in M. tuberculosis H37Rv results from the transfer of Manp from GDP-Manp to polyprenyl phosphates catalysed by the C-terminal domain(s) of a PPM synthase (Mt-Ppm1), now termed as Mt-Ppm1/D2 [22].…”

Section: Introductionmentioning

confidence: 99%

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Novel prenyl-linked benzophenone substrate analogues of mycobacterial mannosyltransferases

Guy

Illarionov

Gurcha

et al. 2004

Biochemical Journal

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PPM (polyprenol monophosphomannose) has been shown to act as a glycosyl donor in the biosynthesis of the Man (mannose)-rich mycobacterial lipoglycans LM (lipomannan) and LAM (lipoarabinomannan). The Mycobacterium tuberculosis PPM synthase (Mt-Ppm1) catalyses the transfer of Man from GDP-Man to polyprenyl phosphates. The resulting PPM then serves as a donor of Man residues leading to the formation of an α(1 → 6)LM intermediate through a PPM-dependent α(1 → 6)mannosyltransferase. In the present study, we prepared a series of ten novel prenyl-related photoactivatable probes based on benzophenone with lipophilic spacers replacing several internal isoprene units. These probes were excellent substrates for the recombinant PPM synthase Mt-Ppm1/D2 and, on photoactivation, several inhibited its activity in vitro. The protection of the PPM synthase activity by a 'natural' C 75 polyprenyl acceptor during phototreatment is consistent with probe-mediated photoinhibition occurring via specific covalent modification of the enzyme active site. In addition, the unique mannosylated derivatives of the photoreactive probes were all donors of Man residues, through a PPM-dependent mycobacterial α(1 → 6)mannosyltransferase, to a synthetic Manp(1 → 6)-Manp-O-C 10:1 disaccharide acceptor (where Manp stands for mannopyranose). Photoactivation of probe 7 led to striking-specific inhibition of the M. smegmatis α(1 → 6)mannosyltransferase. The present study represents the first application of photoreactive probes to the study of mycobacterial glycosyltransferases involved in LM and LAM biosynthesis. These preliminary findings suggest that the probes will prove useful in investigating the polyprenyl-dependent steps of the complex biosynthetic pathways to the mycobacterial lipoglycans, aiding in the identification of novel glycosyltransferases.

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