Autotransporter β-domains have a specific function in protein secretion beyond outer-membrane targeting

Area of Science:

• Microbiology
• Molecular Biology
• Structural Biology

Background:

• Autotransporters (ATs) are Gram-negative bacterial proteins with a passenger domain secreted extracellularly and a C-terminal beta-domain forming an outer membrane (OM) beta-barrel.
• The beta-domain was traditionally viewed as the translocation pore, but its narrow structure contradicts the transport of folded passenger domains.
• Recent findings suggest ATs interact with the beta-barrel assembly machinery, questioning the beta-domain's sole role in translocation.

Purpose of the Study:

• To investigate the precise function of the AT beta-domain in passenger translocation.
• To determine if the beta-domain is merely a targeting device or actively participates in transport.
• To explore the impact of beta-domain modifications on protein secretion.

Main Methods:

• Replaced the native beta-domain of the Escherichia coli AT hemoglobin protease with beta-domains from other OM proteins.
• Engineered mutations to alter the diameter of the beta-domain pore.
• Assessed mutant protein insertion into the OM and passenger display on the cell surface.

Main Results:

• Efficient passenger secretion is dependent on a specific beta-domain.
• The beta-domain acts as a targeting device for the passenger.
• The beta-domain is directly involved in the translocation of the passenger to the cell surface.

Conclusions:

• The beta-domain of autotransporters plays a dual role: targeting the passenger and actively participating in its translocation across the outer membrane.
• The structural and functional specificity of the beta-domain is essential for efficient protein secretion in Gram-negative bacteria.
• This study refines the understanding of the autotransporter mechanism, highlighting the beta-domain's active role beyond simple pore formation.