Characterization of a Mycobacterium tuberculosis nanocompartment and its potential cargo proteins.

Mycobacterium tuberculosis has evolved various mechanisms by which the bacterium can maintain homeostasis under numerous environmental assaults generated by the host immune response. M. tuberculosis harbors enzymes involved in the oxidative stress response that aid in survival during the production of reactive oxygen species in activated macrophages. Previous studies have shown that a dye-decolorizing peroxidase (DyP) is encapsulated by a bacterial nanocompartment, encapsulin (Enc), whereby packaged DyP interacts with Enc via a unique C-terminal extension. M. tuberculosis also harbors an encapsulin homolog (CFP-29, Mt-Enc), within an operon with M. tuberculosis DyP (Mt-DyP), which contains a C-terminal extension. Together these observations suggest that Mt-DyP interacts with Mt-Enc. Furthermore, it has been suggested that DyPs may function as either a heme-dependent peroxidase or a deferrochelatase. Like Mt-DyP, M. tuberculosis iron storage ferritin protein, Mt-BfrB, and an M. tuberculosis protein involved in folate biosynthesis, 7,8-dihydroneopterin aldolase (Mt-FolB), have C-terminal tails that could also interact with Mt-Enc. For the first time, we show by co-purification and electron microscopy that mycobacteria via Mt-Enc can encapsulate Mt-DyP, Mt-BfrB, and Mt-FolB. Functional studies of free or encapsulated proteins demonstrate that they retain their enzymatic activity within the Mt-Enc nanocompartment. Mt-DyP, Mt-FolB, and Mt-BfrB all have antioxidant properties, suggesting that if these proteins are encapsulated by Mt-Enc, then this nanocage may play a role in the M. tuberculosis oxidative stress response. This report provides initial structural and biochemical clues regarding the molecular mechanisms that utilize compartmentalization by which the mycobacterial cell may aid in detoxification of the local environment to ensure long term survival.

Discovery and characterization of a unique mycobacterial heme acquisition system.

Mycobacterium tuberculosis must import iron from its host for survival, and its siderophore-dependent iron acquisition pathways are well established. Here we demonstrate a newly characterized pathway, whereby M. tuberculosis can use free heme and heme from hemoglobin as an iron source. Significantly, we identified the genomic region, Rv0202c-Rv0207c, responsible for the passage of heme iron across the mycobacterial membrane. Key players of this heme uptake system were characterized including a secreted protein and two transmembrane proteins, all three specific to mycobacteria. Furthermore, the crystal structure of the key heme carrier protein Rv0203 was found to have a unique fold. The discovery of a unique mycobacterial heme acquisition pathway opens new avenues of exploration into mycobacterial therapeutics.