Fig. 1: Biosynthetic pathway and structural architecture of PimE.

a Overview of PIM biosynthesis in mycobacteria, featuring the enzymatic reactions carried out by key mannosyltransferases. Enzyme names are indicated above the reaction arrows. The early steps occur on the cytoplasmic side of the plasma membrane, where PimA and PimB transfer mannose residues from GDP-mannose to the 2-OH and 6-OH positions of the inositol moiety of phosphatidylinositol (PI), yielding PIM1 and PIM2, respectively. PatA then acylates the 6-OH position of the mannose ring of PIM2, resulting in AcPIM2. An unknown acyltransferase can add an additional acyl chain to the 3-OH of the inositol of AcPIM2 to form Ac₂PIM2. These acylated mannosylated intermediates are presumed to translocate to the outer leaflet of the membrane by an unknown flippase. PimC and PimD are thought to catalyze the subsequent tri- and tetra-mannosylation steps, leading to the formation of Ac₁PIM4. PimE catalyzes the transfer of the fifth mannose residue from polyprenyl-monophospho-β-d-mannose (PPM) to Ac₁PIM4, forming Ac₁PIM5. The enzyme responsible for the conversion of Ac₁PIM5 to AcPIM6 remains unknown. b Mannosyl transfer reaction catalyzed by PimE, showing the formation of an α(1 → 2) glycosidic bond between PPM and Ac₁PIM4, yielding Ac₁PIM5 and PP. Only tri-acylated PIMs (Ac₁PIM4 and Ac₁PIM5) are shown for the sake of simplicity. c Cryo-EM map of MaPimE in complex with Fab-E6. PimE is depicted in rainbow, while the Fab-E6 is shown in gray. (d) PimE is shown as a ribbon colored in rainbow, as in Fig. 1f. e TM helix arrangement of PimE is depicted as a cross-section colored in a rainbow as in Fig. 1f. f Topological diagram of PimE showing the arrangement of TM segments and extracellular domains. The key catalytic residue D58 is shown as a red dot.