Model of mouth-to-mouth transfer of bacterial lipoproteins through inner membrane LolC, periplasmic LolA, and outer membrane LolB

Abstract

Outer membrane-specific lipoproteins in Escherichia coli are released from the inner membrane by an ATP-binding cassette transporter, the LolCDE complex, which causes the formation of a soluble complex with a periplasmic molecular chaperone, LolA. LolA then transports lipoproteins to the outer membrane where an outer membrane receptor, LolB, incorporates lipoproteins into the outer membrane. The molecular mechanisms underlying the Loldependent lipoprotein sorting have been clarified in detail. However, it remained unclear how Lol factors interact with each other to conduct very efficient lipoprotein transfer in the periplasm where ATP is not available. To address this issue, a photo-reactive phenylalanine analogue, p-benzoyl-phenylalanine, was introduced at various positions of LolA and LolB, of which the overall structures are very similar and comprise an incomplete ␤-barrel with a hydrophobic cavity inside. Cells expressing LolA or LolB derivatives containing the above analogue were irradiated with UV for in vivo photo-cross-linking. These analyses revealed a hot area in the same region of LolA and LolB, through which LolA and LolB interact with each other. This area is located at the entrance of the hydrophobic cavity. Moreover, this area in LolA is involved in the interaction with a membrane subunit, LolC, whereas no cross-linking occurs between LolA and the other membrane subunit, LolE, or ATP-binding subunit LolD, despite the structural similarity between LolC and LolE. The hydrophobic cavities of LolA and LolB were both found to bind lipoproteins inside. These results indicate that the transfer of lipoproteins through Lol proteins occurs in a mouth-to-mouth manner.E. coli ͉ Lol proteins ͉ molecular chaperone ͉ photo-cross-link

Structural Investigation of the Interaction between LolA and LolB Using NMR

“…The photo-cross-linking experiments suggested that some residues that were identified by the preset study exhibited the high efficiency of the cross-linking (28). Therefore, these observations also support the interaction mode.…”

Defective Lipoprotein Sorting Induces lolA Expression through the Rcs Stress Response Phosphorelay System

“…The cavity is covered by a lid comprising short ␣ helices. The inside of the cavity is hydrophobic and binds to the acyl chains of lipoproteins (30). To study the importance of the opening and closing of the lid covering the cavity, we constructed a LolA mutant protein having two mutations, I93C and F140C (43).…”

A Periplasmic LolA Derivative with a Lethal Disulfide Bond Activates the Cpx Stress Response System

“…Membrane localization of LolA(I93C/F140C). It was shown recently by means of in vivo photo-cross-linking that the region around the entrance of the hydrophobic cavity of LolA physically interacts with LolC (20). It therefore seemed likely that the toxic effect of I93C/F140C is suppressed by the formation of the I93C/F140C-LolCDE complex.…”

“…Taken together, these results indicate that the oxidized form of I93C/F140C more strongly binds to LolCDE. It was recently found by means of an in vivo photo-cross-linking technique that LolA specifically interacts with LolC but not LolE (20), despite their similar structures (37). It therefore seems likely that I93C/F140C binds to the LolC subunit of LolCDE.…”

“…LolA, a periplasmic lipoprotein-specific carrier, receives lipoproteins from LolCDE in an ATP hydrolysis-dependent manner and forms a water-soluble complex with a lipoprotein (13). The complex then traverses the periplasmic space from the inner to the outer membrane, where lipoproteins are transferred from LolA to a lipoprotein receptor, LolB (14), in a mouth-to-mouth manner (20). Finally, lipoproteins are anchored to the outer membrane through the action of LolB (32).…”