Substrate binding to BamD triggers a conformational change in BamA to control membrane insertion

Abstract

<p id="d7612089e363">The outer membrane of Gram-negative bacteria, mitochondria, and chloroplasts contains proteins that adopt β-barrel structures. To maintain the integrity of this structure, the β-barrel assembly machine (Bam) folds and inserts integral membrane proteins into the outer membranes of Gram-negative bacteria. By studying the assembly of an essential β-barrel substrate we found that the two essential components of the complex, BamA and BamD, coordinate with each other upon substrate binding to facilitate folding and insertion. These results could enable the design of strategies to combat Gram-negative pathogens. </p><p class="first" id="d7612089e366">The β-barrel assembly machine (Bam) complex folds and inserts integral membrane proteins into the outer membrane of Gram-negative bacteria. The two essential components of the complex, BamA and BamD, both interact with substrates, but how the two coordinate with each other during assembly is not clear. To elucidate aspects of this process we slowed the assembly of an essential β-barrel substrate of the Bam complex, LptD, by changing a conserved residue near the C terminus. This defective substrate is recruited to the Bam complex via BamD but is unable to integrate into the membrane efficiently. Changes in the extracellular loops of BamA partially restore assembly kinetics, implying that BamA fails to engage this defective substrate. We conclude that substrate binding to BamD activates BamA by regulating extracellular loop interactions for folding and membrane integration. </p>