Folding kinetics of the outer membrane proteins OmpA and FomA into phospholipid bilayers

Abstract

The folding mechanism of outer membrane proteins (OMPs) of Gram-negative bacteria into lipid bilayers has been studied using OmpA of E. coli and FomA of F. nucleatum as examples. Both, OmpA and FomA are soluble in unfolded form in urea and insert and fold into phospholipid bilayers upon strong dilution of the denaturant urea. OmpA is a structural protein and forms a small ion channel, composed of an 8-stranded transmembrane ␤-barrel domain. FomA is a voltage-dependent porin, predicted to form a 14 stranded ␤-barrel. Both OMPs fold into a range of model membranes of very different phospholipid compositions. Three membrane-bound folding intermediates of OmpA were discovered in folding studies with dioleoylphosphatidylcholine bilayers that demonstrated a highly synchronized mechanism of secondary and tertiary structure formation of ␤-barrel membrane proteins. A study on FomA folding into lipid bilayers indicated the presence of parallel folding pathways for OMPs with larger transmembrane ␤-barrels.

Correct Folding of the β-Barrel of the Human Membrane Protein VDAC Requires a Lipid Bilayer

“…Kleinschmidt 3 and references therein). Folded, solubilized and unfolded forms of hVDAC1 did not show any differences in electrophoretic mobility, neither in SDS-PAGE nor in native gel 25 analysis.…”

“…Folded OMPs of bacteria are usually stable in SDS at RT and migrate differently when not heat-denatured prior to SDS-PAGE (see e.g. Kleinschmidt for a review 3 ). This was not observed for hVDAC1 as a consequence of the altered secondary (and tertiary) structure in SDS.…”

“…reviews 1,2 and references therein) or with outer membrane protein A (OmpA) of Escherichia coli (see e.g. review 3 and references therein), which forms a TM β-barrel domain. BR and OmpA belong to the two known classes of TMPs that are distinguished by their TM secondary structure.…”

“…Analysis II was performed on hVDAC1 that was first solubilized in LDAO and then reconstituted into diC 12:0 PC (the CD spectrum is shown in Figure 1 3,5,7,9,12,14,16), and in diC 12:0 PC (lanes 4,6,8,10,13,15,17), indicate formation of a single fragment of ∼ 26 to 27 kDa size in diC 12:0 PC bilayers that is present after 240 min of proteolysis. In LDAO, hVDAC1 is first cleaved into two major fragments of ∼ 26 to 27 kDa and of ∼ 24 kDa.…”

VirK Is a Periplasmic Protein Required for Efficient Secretion of Plasmid-Encoded Toxin from Enteroaggregative Escherichia coli

“…In recent years, several studies have demonstrated that periplasmic proteins can serve as molecular chaperones in the assembly pathway of OMPs. For example, deletion of the genes encoding the 17-kDa Skp protein or the 48-kDa protein survival factor A (SurA) resulted in reduced concentrations of OMPs in the OM (17). Skp, SurA, and DegP are parts of a functional network that is vital for proper protein folding and degradation in the cell envelope (29).…”