Characterization of phthiocerol and phthiodiolone dimycocerosate esters of M. tuberculosis by multiple-stage linear ion-trap MS

Abstract

This article is available online at http://www.jlr.org Both phthiocerol/phthiodiolone dimycocerosate (PDIM) esters and phenolic glycolipids (PGLs) are dimycocerosate esters (DIMs) produced by pathogenic mycobacteria. PDIMs were originally isolated from Mycobacterium tuberculosis (1)(2)(3)(4) and are specifi c tuberculosis biomarkers ( 5, 6 ). They are among the most abundant lipids in the cell wall of various pathogenic mycobacteria, including M. tuberculosis , M. bovis , M. leprae , M. kansasii , M. microti , and M. marinum ( 7-10 ), which are known to synthesize a range of complex highmolecular-mass lipids ( 7 ). PGLs are produced by the same set of pathogenic mycobacteria species, except that in M. tuberculosis only a subset of clinical isolates belonging to the W-Beijing family ( 11 ) produces PGLs.Phthiocerol and phenolphthiocerol and other lipids such as mycolic acids and methyl-branched FAs in cell wall are among those that have been most extensively studied in terms of their biosynthesis and the role in M. tuberculosis virulence in vivo ( 12 ). In pathogenesis, the role of PDIMs of M. tuberculosis was recognized by the studies that identifi ed that mutants of M. tuberculosis were unable to either produce or properly localize PDIMs to the cell envelopeand that demonstrated that PDIM-defi cient strains were attenuated in animal models of infection ( 8,11,(13)(14)(15)(16)(17). Recently, M. tuberculosis and its close pathogenic relative M. marinum were reported to manipulate macrophage recruitment through coordinated use of membrane PDIM and PGLs to initiate infections ( 18 ). However, the precise role of these molecules in the course of infection remains largely unknown, and their role in the multiplication of mycobacteria from the tuberculosis complex in organs other than the lungs is also unclear.

Mycobacterium tuberculosis carrying a rifampicin drug resistance mutation reprograms macrophage metabolism through cell wall lipid changes

“…11. The normalized spectra were presented in Figure 3, where the ESI/MS spectra contain the homologous [M+NH 4 ] + ions of PDIM, ranging from m/z 1250 to 1550 (i.e., ions of m/z 1343, 1357, 1371, 1385, 1399, 1413, etc) 49 .…”

Multiple-stage Precursor Ion Separation and High Resolution Mass Spectrometry toward Structural Characterization of 2,3-Diacyltrehalose Family from Mycobacterium tuberculosis

“…For example, we demonstrated that sulfolipid II in Mycobacterium tuberculosis (M. tuberculosis) H37Rv cells is the predominated lipid family, consisting of hundreds of molecular species rather than the sulfolipid I family as previously reported [8,9]. LIT MS n with high resolution mass spectrometry has also been successfully applied to delineate the structures of phosphatidylinositol mannosides (PIMs) [10,11], and phthiocerol dimycocerosates (PDIMs) [12] in the cell envelope of M. tuberculosis. The former lipid family is known to have played important roles in M. tuberculosis adhesins that mediate attachment to nonphagocytic cells [13], while the latter is recently found to play role in drug resistance to M. tuberculosis [14].…”

“…tuberculosis strain H37Rv were grown and total lipids were extracted and isolated as previously described [8]. Briefly, the total lipid was separated by a Phenomenex C18 Kinetex (100 × 4.6 mm, pore size 100 Å, particle size 2.6 µm) column at a flow rate of 300 µL/min with a gradient system as previously described [12]. DAT (eluted at 24.45-26.43 min) fraction from three injections (~200 µg total lipid/injection) were collected and pooled, dried under a stream of nitrogen, and stored at −20 • C until use.…”

Roles for phthiocerol dimycocerosate lipids in Mycobacterium tuberculosis pathogenesis

“…Thus, using solid media for passages may allow the preservation of PDIM positive strains. Dedicated methods to test lab strains for the presence of PDIMs are thin-layer chromatography and mass spectrometry, both of which can be coupled with NMR spectroscopy for complete characterization [48,116]. For an easier and relatively faster routine procedure, lab strains could be tested indirectly for the presence of PDIMs by drug-susceptibility assay, using drugs shown to be more potent on PDIM-depleted strains such as vancomycin [55].…”