title: Scrutiny of Mycobacterium tuberculosis 19 kDa antigen proteoforms provides new insights in the lipoglycoprotein biogenesis paradigm.
abstract: Post-translational modifications (PTMs) are essential processes conditioning the biophysical properties and biological activities of the vast majority of mature proteins. However, occurrence of several distinct PTMs on a same protein dramatically increases its molecular diversity. The comprehensive understanding of the functionalities resulting from any particular PTM association requires a highly challenging full structural description of the PTM combinations. Here, we report the in-depth exploration of the natural structural diversity of the M. tuberculosis (Mtb) virulence associated 19 kDa lipoglycoprotein antigen (LpqH) using intact protein high-resolution mass spectrometry (HR-MS) coupled to liquid chromatography. Combined top-down and bottom-up HR-MS analyses of the purified Mtb LpqH protein allow, for the first time, to uncover a complex repertoire of about 130 molecular species resulting from the intrinsically heterogeneous combination of lipidation and glycosylation together with some truncations. Direct view on the co-occurring PTMs stoichiometry reveals the presence of functionally distinct LpqH lipidation states and indicates that glycosylation is independent from lipidation. This work allowed the identification of a novel unsuspected phosphorylated form of the unprocessed preprolipoglycoprotein totally absent from the current lipoglycoprotein biogenesis pathway and providing new insights into the biogenesis and functional determinants of the mycobacterial lipoglycoprotein interacting with the host immune PRRs.
authors.affiliation: Institut de Pharmacologie et de Biologie Structurale, Université de Toulouse, CNRS, UPS, France.
authors.forename: Julien
authors.initials: J
authors.lastname: Parra
authors.affiliation: Institut de Pharmacologie et de Biologie Structurale, Université de Toulouse, CNRS, UPS, France.
authors.forename: Julien
authors.initials: J
authors.lastname: Marcoux
authors.affiliation: Aix Marseille Univ, CNRS, EIPL, IMM FR3479, Marseille, France.
authors.forename: Isabelle
authors.initials: I
authors.lastname: Poncin
authors.affiliation: Aix Marseille Univ, CNRS, EIPL, IMM FR3479, Marseille, France.
authors.forename: Stéphane
authors.initials: S
authors.lastname: Canaan
authors.affiliation: UMR1173, INSERM, Université de Versailles St Quentin, 78180, Montigny le Bretonneux, France.
authors.forename: Jean Louis
authors.initials: JL
authors.lastname: Herrmann
authors.affiliation: Institut de Pharmacologie et de Biologie Structurale, Université de Toulouse, CNRS, UPS, France.
authors.forename: Jérôme
authors.initials: J
authors.lastname: Nigou
authors.affiliation: Institut de Pharmacologie et de Biologie Structurale, Université de Toulouse, CNRS, UPS, France.
authors.forename: Odile
authors.initials: O
authors.lastname: Burlet-Schiltz
authors.affiliation: Institut de Pharmacologie et de Biologie Structurale, Université de Toulouse, CNRS, UPS, France.
authors.forename: Michel
authors.initials: M
authors.lastname: Rivière
mesh_terms: D000107:Acetylation; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000276:immunology / Q000378:metabolism*; D002853:Chromatography, Liquid; D006023:Glycoproteins / Q000096:biosynthesis*; D008074:Lipoproteins / Q000096:biosynthesis*; D008954:Models, Biological; D009169:Mycobacterium tuberculosis / Q000276:immunology / Q000378:metabolism*; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D020543:Proteome; D040901:Proteomics / Q000379:methods; D053719:Tandem Mass Spectrometry
chemical_list: C087864:19 kDa antigen, Mycobacterium; D001426:Bacterial Proteins; D006023:Glycoproteins; D008074:Lipoproteins; D020543:Proteome