Structural Insights on the Mycobacterium tuberculosis Proteasomal ATPase Mpa

Biology Department, Brookhaven National Laboratory, Upton, NY 11973-5000, USA.
Structure (Impact Factor: 5.62). 10/2009; 17(10):1377-85. DOI: 10.1016/j.str.2009.08.010
Source: PubMed


Proteasome-mediated protein turnover in all domains of life is an energy-dependent process that requires ATPase activity. Mycobacterium tuberculosis (Mtb) was recently shown to possess a ubiquitin-like proteasome pathway that plays an essential role in Mtb resistance to killing by products of host macrophages. Here we report our structural and biochemical investigation of Mpa, the presumptive Mtb proteasomal ATPase. We demonstrate that Mpa binds to the Mtb proteasome in the presence of ATPgammaS, providing the physical evidence that Mpa is the proteasomal ATPase. X-ray crystallographic determination of the conserved interdomain showed a five stranded double beta barrel structure containing a Greek key motif. Structure and mutational analysis indicate a major role of the interdomain for Mpa hexamerization. Our mutational and functional studies further suggest that the central channel in the Mpa hexamer is involved in protein substrate translocation and degradation. These studies provide insights into how a bacterial proteasomal ATPase interacts with and facilitates protein degradation by the proteasome.

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    • "In this work STINT-NMR was used to examine the interactions between the Ubiquitin-like protein, Pup, in the presence of the Mycobacterium proteasome ATPase, Mpa, and the active 1.2 megadalton proteasome complex, consisting of Mpa and the Mycobacterial proteasome core particle (CP) [9,10]. The importance of this macromolecular complex is underlined by the fact that Mycobacterium tuberculosis is particularly resistant to reactive nitrogen intermediates generated by host immune system, and this resistance is related to the proteasome [11] and mpa [12]. "
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    ABSTRACT: The Mycobacterium tuberculosis proteasome is required for maximum virulence and to resist killing by the host immune system. The prokaryotic ubiquitin-like protein, Pup-GGE, targets proteins for proteasome-mediated degradation. We demonstrate that Pup-GGQ, a precursor of Pup-GGE, is not a substrate for proteasomal degradation. Using STINT-NMR, an in-cell NMR technique, we studied the interactions between Pup-GGQ, mycobacterial proteasomal ATPase, Mpa, and Mtb proteasome core particle (CP) inside a living cell at amino acid residue resolution. We showed that under in-cell conditions, in the absence of the proteasome CP, Pup-GGQ interacts with Mpa only weakly, primarily through its C-terminal region. When Mpa and non-stoichiometric amounts of proteasome CP are present, both the N-terminal and C-terminal regions of Pup-GGQ bind strongly to Mpa. This suggests a mechanism by which transient binding of Mpa to the proteasome CP controls the fate of Pup.
    Full-text · Article · Sep 2013 · PLoS ONE
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    • "In Mtb, two putative accessory factors PafA and Mpa are involved in coupling the prokaryotic ubiquitin-like protein Pup to peptides that are destined for turnover and recognition of Pup-tagged proteins, respectively (Pearce et al., 2008; Striebel et al., 2009; Sutter et al., 2009). Mpa is an ATPase likely involved in unfolding and delivery of proteins into the proteolytic core of proteasomes (Wang et al., 2009). Mutations in the respective genes mpa and pafA increase the sensitivity of the tubercle bacillus to RNS (Darwin, 2009). "
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    ABSTRACT: Tuberculosis (TB) remains a major health threat, killing nearly 2 million individuals around this globe, annually. The only vaccine, developed almost a century ago, provides limited protection only during childhood. After decades without the introduction of new antibiotics, several candidates are currently undergoing clinical investigation. Curing TB requires prolonged combination of chemotherapy with several drugs. Moreover, monitoring the success of therapy is questionable owing to the lack of reliable biomarkers. To substantially improve the situation, a detailed understanding of the cross-talk between human host and the pathogen Mycobacterium tuberculosis (Mtb) is vital. Principally, the enormous success of Mtb is based on three capacities: first, reprogramming of macrophages after primary infection/phagocytosis to prevent its own destruction; second, initiating the formation of well-organized granulomas, comprising different immune cells to create a confined environment for the host-pathogen standoff; third, the capability to shut down its own central metabolism, terminate replication, and thereby transit into a stage of dormancy rendering itself extremely resistant to host defense and drug treatment. Here, we review the molecular mechanisms underlying these processes, draw conclusions in a working model of mycobacterial dormancy, and highlight gaps in our understanding to be addressed in future research.
    Preview · Article · Feb 2012 · FEMS microbiology reviews
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    • "Biochemical studies with Mpa and ARC (AAA ATPase forming ring-shaped complexes), an orthologous ATPase from Rhodococcus erythropolis, showed that the ATPases formed hexameric or dodecameric rings with ATPase activity (Wolf et al., 1998, Zhang et al., 2004, Darwin et al., 2005). It is hypothesized that Mpa and ARC perform an analogous function to the AAA ATPases of the eukaryotic RP, however, robust interactions between the bacterial ATPases and CPs have not been observed (Wolf et al., 1998, Wang et al., 2009). This suggests that the interactions are either transient or require additional factors. "
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    ABSTRACT: Prokaryotic ubiquitin-like protein (Pup) is the first identified prokaryotic protein that is functionally analogous to ubiquitin. Despite using the proteasome as the end-point for proteolysis, Pup differs from ubiquitin both biochemically and structurally. We will discuss these differences that have been highlighted by several recent studies. Finally, we will speculate on the possible interactions between the two analogous pathways in pathogen and host.
    Preview · Article · Apr 2010 · Cellular Microbiology
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