Proteasome substrate capture and gate opening by the accessory factor PafE from Mycobacterium tuberculosis

Area of Science:

• Molecular Biology
• Microbiology
• Structural Biology

Background:

• Proteasomes are essential cellular machines for protein degradation, requiring activators to open their gates.
• Mycobacterium tuberculosis virulence depends on proteasomal degradation, facilitated by proteasome accessory factor E (PafE).
• Unlike known activators, PafE functions independently of ATP.

Purpose of the Study:

• To elucidate the mechanism of PafE-mediated protein targeting and proteasome activation.
• To identify PafE interactions with proteasome core particles and native substrates.
• To characterize the role of the conserved GQYL motif in PafE function.

Main Methods:

• Cryo-electron microscopy (cryo-EM) to determine structural interactions.
• Biochemical assays to study proteolysis and substrate binding.
• Identification and characterization of novel proteasome substrates.

Main Results:

• The conserved GQYL motif in PafE directly interacts with proteasome α subunits, triggering gate opening.
• PafE forms dodecameric rings with 40-Å openings, creating a chamber for substrate capture.
• A novel substrate, Rv3213c, was identified and shown to be degraded via the PafE pathway.
• PafE exhibits previously unrecognized chaperone activity in substrate capture.

Conclusions:

• PafE utilizes its GQYL motif to interact with the proteasome core particle, inducing conformational changes for gate opening.
• PafE possesses a dual function: proteasome activation and substrate capture, indicating a novel chaperone-like activity.
• This study reveals the mechanism of ATP-independent proteasome activation and degradation in bacteria.