Ajay K. Mahalka - Effect of multi-drug resistance ABC transporter activity...
Channel: Institut des Hautes Études Scientifiques (IHÉS)
Date: 2018-03-26

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
, ajay.mahalka@curie.fr*

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