Dynamic topology-mediated maturation of β-barrel proteins in BAM-catalyzed folding

Abstract

Abstract BamA is an essential component of the β-barrel assembly complex (BAM) that catalyzes the insertion and assembly of β-barrel outer membrane proteins (OMPs) in Gram-negative bacteria. BamA adopts two distinct conformations: the inward-open and outward-open β-barrels, regulated by periplasmic domains during different folding stages. However, it remains unclear how these conformational shifts promote the maturation of OMPs. Drawing from our prior work1, we identify a novel folding intermediate, characterized by an inward-open BamA and a non-closing OMP. This intermediate attains stability via enhanced hydrophobic interactions accompanying the topological alterations of the hybrid-barrel. The subsequent folding events of the hybrid-barrel are then sampled through molecular dynamics simulations augmented with enhanced sampling. Remarkably, our observations reveal that the spontaneous closure and separation are regulated by an induced-fit mechanism and supplementary hydrophobic interactions, prominently from the extracellular loop 1 (BL1) of BamA. Further cell viability assays confirm that disruption to BL1’s topology compromises BamA functionality and is detrimental to the bacteria. Taken together, our findings provide insights into the significance of BamA’s dynamic topological shifts during OMP maturation.