Mechanistic insights into TAM-mediated OMP assembly in Gram-negative bacteria

Area of Science:

• Microbiology
• Structural Biology
• Biochemistry

Background:

• Outer membrane protein (OMP) biogenesis is crucial for Gram-negative bacteria.
• The β-barrel assembly machinery (BAM) is essential for OMP assembly and a target for antibiotics.
• The translocation and assembly module (TAM) is an evolutionary relative of BAM, but its mechanism is poorly understood.

Purpose of the Study:

• To elucidate the mechanism of OMP assembly by the TAM complex.
• To determine the structures of key TAM components, TamA and TamAB, in various conformational states.

Main Methods:

• Cryo-electron microscopy (cryo-EM) was used to obtain high-resolution structures of Escherichia coli TamA and TamAB.
• NanoLuc Binary Technology (NanoBiT)-based folding assays were employed to assess OMP assembly efficiency.

Main Results:

• Cryo-EM structures revealed multiple conformational states of TamA and TamAB.
• The TamB C-terminal tail was shown to engage the TamA β-barrel, opening its lateral gate and forming a hybrid β-interface.
• An extended AsmA-like periplasmic conduit was activated, potentially facilitating substrate transport.
• TamAB demonstrated more efficient OMP assembly than TamA alone, dependent on lateral-gate opening and hybrid-interface formation.

Conclusions:

• A substrate-mimetic priming mechanism for OMP biogenesis by TAM was defined, distinguishing it from BAM.
• The study establishes a structural framework for understanding TAM-mediated OMP biogenesis.