Ajay K. Mahalka - Effect of multi-drug resistance ABC transporter activity...

Effect of multi-drug resistance ABC transporter activity on their conformation-induced redistribution on membranes

Effect of multi-drug resistance ABC transporter activity on their
conformation-induced redistribution on membranes
Ajay K. Mahalka1*, Paik Su-Jin2,4, John Manzi3,Daniel Levy2, and Patricia Bassereau11
Membranes and cellular functions, 2Molecular Microscopy of Membranes, 4 Biochemistry and Molecular Biology Platform, Institut Curie, UMR 168 and 3 PSL Research University Paris, France
, [email protected]*

Integral trans-membrane proteins are involved in various cellular functions and their dysfunction is associated with human pathologies [1]. The lipid-protein interactions have been studied to address structure-function relationship of transmembrane proteins at molecular level. However, the effects related to membranes physical properties on trans-membrane proteins have not been well-studied, and not at all when their conformations change. Recent experimental evidence indicates the intrinsic interplay between protein shape and the properties of its membrane environment [2,3]. It is expected that non-cylindrical proteins tend to cluster and be enriched in curved membranes. Thus, we studied BmrA a bacterial ATP binding cassette (ABC) transporter from B. subtilisinvolved in export of a large diversity of substrates in an ATP dependent manner,
fairly homologous to human P-glycoprotein ,[4]. The conformational change in nucleotide-binding domains (NBDs) of
BmrA between apo and the post-hydrolytic state (tweezers-like
motion) is 5 nm and that is the largest tweezers motion
reported till date in the case of trans-membrane proteins.
Here we addressed how the conformational dynamics of BmrA influence its membrane properties, in particular its spatial distribution on flat or curved membranes.

To decipher the effect of the conformational dynamics of Bm
rA on its spatial distribution in membranes,
depending on membrane curvature, we used cell-sized giant unilamellar vesicles (GUVs) containing either the apo-or closed-conformation BmrA to form membrane nanotubes with controlled radii. We found that, at low protein density, apo-BmrA
is highly enriched (50 times) in nanotubes as compared to flat membrane and simultaneously modulates tube radius from 100 nm to 30 nm, due to its high intrinsic curvature. Surprisingly, although the post-hydrolytic closed-conformation BmrA is expected to be cylindrical, we measured an enrichment
of this conformation in nanotubes, but about 3 time less
pronounced then for apo-BmrA. Eventually, in the presence of ATP, BmrAhas reduced curvature selectivity as compared to the apo form, in agreement with a cycling change of conformation between the apo and the closed forms. This study on reconstituted transmembrane proteins demonstrates that protein distribution on membranes is influenced by the interplay of membrane curvature, effective shape and flexibility of membrane proteins.

[1] Gottesman M. et al., (2002) Multidrug resistance in cancer: role of ATP-dependent transporters. Nat Rev Cancer 2: 48-58
[2] Quemeneur F. et al.,(2014) Shape matters in protein mobility within membranes. PNAS 111:5083-5087
[3] Aimon S. et al., (2014) Membrane shape modulates trans-membrane protein distribution. Dev Cell 28: 212-218.
[4] Lubelski J.et al., (2007) Distribution and Physiology of ABC-Type Transporters Contributing to Multidrug Resistance in Bacteria. Microbiol MolBiol Rev 71: 463-476