Repacking of the transmembrane domains of P-glycoprotein during the transport ATPase cycle

Rosenberg, MF; Velarde, G; Ford, RC; Martin, C; Berridge, G; Kerr, ID; Callaghan, R; Schmidlin, A; Wooding, C; Linton, KJ; Higgins, CF

Abstract

P-glycoprotein (P-gp) is an ABC (ATP-binding cassette) transporter, which hydrolyses ATP and extrudes cytotoxic drugs from mammalian cells. P-gp consists of two transmembrane domains (TMDs) that span the membrane multiple times, and two cytoplasmic nucleotide-binding domains (NBDs). We have determined projection structures of P-gp trapped at different steps of the transport cycle and correlated these structures with function. In the absence of nucleotide, an 10 Å resolution structure was determined by electron cryo-microscopy of two-dimensional crystals. The TMDs form a chamber within the membrane that appears to be open to the extracellular milieu, and may also be accessible from the lipid phase at the interfaces between the two TMDs. Nucleotide binding causes a repacking of the TMDs and reduction in drug binding affinity. Thus, ATP binding, not hydrolysis, drives the major conformational change associated with solute translocation. A third distinct conformation of the protein was observed in the post-hydrolytic transition state prior to release of ADP/Pi. Biochemical data suggest that these rearrangements may involve rotation of transmembrane -helices. A mechanism for transport is suggested.

Citation

Rosenberg, M., Velarde, G., Ford, R., Martin, C., Berridge, G., Kerr, I., …Higgins, C. (2001). Repacking of the transmembrane domains of P-glycoprotein during the transport ATPase cycle. The EMBO Journal, 20(20), 5615-5625. https://doi.org/10.1093/emboj/20.20.5615

• Journal Article Type: Article
• Publication Date: Oct 1, 2001
• Deposit Date: May 15, 2007
• Journal: EMBO Journal
• Print ISSN: 0261-4189
• Electronic ISSN: 1460-2075
• Publisher: EMBO Press
• Peer Reviewed: Not Peer Reviewed
• Volume: 20
• Issue: 20
• Pages: 5615-5625
• DOI: https://doi.org/10.1093/emboj/20.20.5615
• Keywords: ABC transporters, Membrane protein structure, Multidrug resistance, Two-dimensional crystals.
• Public URL: https://durham-repository.worktribe.com/output/1577488