FkpA enhances membrane protein folding using an extensive interaction surface.

Area of Science:

• Microbiology
• Molecular Biology
• Biochemistry

Background:

• Gram-negative bacteria rely on periplasmic chaperones like SurA, Skp, and FkpA for outer membrane protein (OMP) biogenesis.
• These chaperones prevent aggregation and facilitate folding of unfolded OMPs (uOMPs).
• FkpA's specific role in OMP folding, particularly under heat-shock stress, is less understood compared to other chaperones.

Purpose of the Study:

• To elucidate the chaperone function of FkpA in the context of OMP folding.
• To investigate how FkpA influences the folding rate and yield of OMPs.
• To characterize the binding interaction between FkpA and unfolded OMPs.

Main Methods:

• Monitoring the folding of three OMPs under FkpA influence.
• Sedimentation velocity (SV) experiments to determine binding affinity and complex formation.
• Photo-crosslinking experiments to map the FkpA-uOMP binding interface.
• Utilizing subdomain constructs of FkpA to assess the role of its full length.

Main Results:

• FkpA increases the folded yield but decreases the folding rate of OMPs, acting as a chaperone rather than a catalyst.
• FkpA binds to all three tested unfolded OMPs with an affinity intermediate between Skp and SurA.
• Complex formation is highly dependent on urea concentration, indicating an extensive binding interface spanning the entire FkpA molecule.
• Full-length FkpA is essential for its complete chaperone activity.

Conclusions:

• FkpA exerts a distinct and direct influence on OMP folding trajectory.
• The chaperone activity of FkpA is mediated through a broad interaction interface with its clients.
• Understanding FkpA's mechanism provides insights into the complex network of OMP biogenesis in bacteria.