Outer membrane protein assembly
The outer membranes of Gram-negative bacteria, mitochondria, and chloroplasts contain beta-barrel integral membrane proteins. Although the folded, membrane-integrated form of a beta-barrel is at a thermodynamic minimum, conserved protein complexes exist that catalyze the folding process. Our lab identified the five-membered protein complex, named Bam (beta-barrel assembly machine), that assembles outer membrane beta-barrel proteins in Gram-negative bacteria.
Outer membrane protein biogenesis
Outer membrane beta-barrels are folded and inserted into the outer membrane by the Bam complex (BamABCDE).
We have purified the protein components of the Bam complex and reconstituted beta-barrel assembly activity in vitro. However, the mechanism by which the Bam complex binds, folds, and inserts its substrates into the outer membrane remains unclear. Our lab uses in vitro and in vivo biochemical tools to characterize the molecular mechanism of beta-barrel assembly. One method we use is the generation of substrates that stall during their assembly, which allows us to probe their interactions with the Bam complex. Our long-term goal is to use mechanistic information we obtain to develop ways of inhibiting the Bam complex as a means of targeting pathogenic Gram-negative bacteria.
Substrate binding to BamD triggers a conformational change in BamA to control membrane insertion. J. Lee, H.A. Sutterlin, J.S. Wzorek, M.D. Mandler, C.L. Hagan, M. Grabowicz, D. Tomasek, M.D. May, E.M. Hart, T.J. Silhavy, D. Kahne. Proc Natl Acad Sci USA 2018; 115:2359-64.
Membrane integration of an essential beta-barrel protein prerequires burial of an extracellular loop. J.S. Wzorek, J. Lee, D. Tomasek, C.L. Hagan, D.E. Kahne. Proc Natl Acad Sci USA 2017; 114:2598-2603.
Characterization of a stalled complex on the beta-barrel assembly machine. J. Lee, M. Xue, J.S. Wzorek, T. Wu, M. Grabowicz, L.S. Gronenberg, H.A. Sutterlin, R.M. Davis, N. Ruiz, T.J. Silhavy, D.E. Kahne. Proc Natl Acad Sci USA 2016; 113:8717-22.
Inhibition of the beta-barrel assembly machine by a peptide that binds BamD. C.L. Hagan, J.S. Wzorek, D. Kahne. Proc Natl Acad Sci USA 2015; 112:2011-6.
Disulfide rearrangement triggered by translocon assembly controls lipopolysaccharide export. S.S. Chng, M. Xue, R.A. Garner, H. Kadokura, D. Boyd, J. Beckwith, D. Kahne. Science 2012; 337:1665-8.
Reconstitution of outer membrane protein assembly from purified components. C.L. Hagan, S. Kim, D. Kahne. Science 2010; 328:890-2.