The Lipoprotein BamB Facilitates Folding and Insertion of Outer Membrane Protein A (OmpA) into Lipid Membranes Depending on Bilayer Thickness

Abstract

Insertion and folding of β-barrel membrane proteins (β-MPs) into lipid bilayers was investigated for outer membrane protein A (OmpA). In Escherichia coli, the assembly of β-MPs is mediated by the barrel assembly machinery (BAM) complex. This complex is composed of the β-MP BamA and 4 lipoproteins, BamB, C, D, and E. We have previously examined the impact of individual BAM proteins, like BamA on insertion and folding of OmpA [1]. Here, BamB was isolated from membrane fractions of E. coli to investigate its properties. BamB folding was reversible even in the absence of lipids as examined by circular dichroism spectroscopy. BamB bound to preformed lipid bilayers of DLPC at a stoichiometry of ∼580±125. The effects of BamB alone on the kinetics of folding and insertion of OmpA were explored for lipid bilayers of different thickness. Interestingly, BamB improved the rates and yields of folding of OmpA into lipid bilayers of DLPC/DLPE/DLPG (5:3:2), but not into lipid bilayers of DOPC/DOPE/DOPG (5:3:2). This was observed for peripherally bound BamB, but not for cytochrome c, which was used for comparisons. A thorough analysis of the folding kinetics of OmpA suggests a stoichiometry of 1:1 for transient interactions of OmpA with BamB. [1] G. Patel, J.H. Kleinschmidt, Biochemistry, 52 (2013) 3974-3986.