Crystal structure of CmABCB1 multi-drug exporter in lipidic mesophase revealed by LCP-SFX

• Pan, Dongqing

  Department of Structural Biology, Graduate School of Pharmaceutical Sciences, Kyoto University

• Oyama, Ryo

  Department of Structural Biology, Graduate School of Pharmaceutical Sciences, Kyoto University

• Sato, Tomomi

  Department of Structural Biology, Graduate School of Pharmaceutical Sciences, Kyoto University

• Nakane, Takanori

  Department of Biological Science, Graduate School of Science, The University of Tokyo

• Mizunuma, Ryo

  Department of Structural Biology, Graduate School of Pharmaceutical Sciences, Kyoto University

• Matsuoka, Keita

  Department of Structural Biology, Graduate School of Pharmaceutical Sciences, Kyoto University

• Joti, Yasumasa

  Japan Synchrotron Radiation Research Institute

• Tono, Kensuke

  Japan Synchrotron Radiation Research Institute

• Nango, Eriko

  RIKEN SPring-8 Center

• Iwata, So

  RIKEN SPring-8 Center; Department of Cell Biology, Graduate School of Medicine, Kyoto University

• Nakatsu, Toru

  Department of Structural Biology, Graduate School of Pharmaceutical Sciences, Kyoto University; RIKEN SPring-8 Center

• Kato, Hiroaki

  Department of Structural Biology, Graduate School of Pharmaceutical Sciences, Kyoto University; RIKEN SPring-8 Center

Abstract

CmABCB1 is a Cyanidioschyzon merolae homolog of human ABCB1, a well known ATP-binding cassette (ABC) transporter responsible for multi-drug resistance in various cancers. Three-dimensional structures of ABCB1 homologs have revealed the snapshots of inward- and outward-facing states of the transporters in action. However, sufficient information to establish the sequential movements of the open–close cycles of the alternating-access model is still lacking. Serial femtosecond crystallography (SFX) using X-ray free-electron lasers has proven its worth in determining novel structures and recording sequential conformational changes of proteins at room temperature, especially for medically important membrane proteins, but it has never been applied to ABC transporters. In this study, 7.7 mono­acyl­glycerol with cholesterol as the host lipid was used and obtained well diffracting microcrystals of the 130 kDa CmABCB1 dimer. Successful SFX experiments were performed by adjusting the viscosity of the crystal suspension of the sponge phase with hy­droxy­propyl methyl­cellulose and using the high-viscosity sample injector for data collection at the SACLA beamline. An outward-facing structure of CmABCB1 at a maximum resolution of 2.22 Å is reported, determined by SFX experiments with crystals formed in the lipidic cubic phase (LCP-SFX), which has never been applied to ABC transporters. In the type I crystal, CmABCB1 dimers interact with adjacent molecules via not only the nucleotide-binding domains but also the transmembrane domains (TMDs); such an interaction was not observed in the previous type II crystal. Although most parts of the structure are similar to those in the previous type II structure, the substrate-exit region of the TMD adopts a different configuration in the type I structure. This difference between the two types of structures reflects the flexibility of the substrate-exit region of CmABCB1, which might be essential for the smooth release of various substrates from the transporter.

Journal

• IUCrJ

  IUCrJ 9 (1), 134-145, 2022-01

  International Union of Crystallography (IUCr)

Keywords

• serial crystallography
• SFX
• protein structures
• sample delivery
• XFELs
• LCP
• lipidic mesophase
• multi-drug exporters
• ABC transporters
• Cyanidioschyzon merolae
• CmABCB1

Details

• CRID: 1050572012354796288
• NII Article ID: 120007179308
• DOI: 10.1107/s2052252521011611
• ISSN: 20522525
• HANDLE: 2433/266705
• Web Site: http://journals.iucr.org/m/issues/2022/01/00/cw5035/cw5035.pdf
• Text Lang: en
• Article Type: journal article
• Data Source:
  • IRDB
  • Crossref
  • CiNii Articles
  • KAKEN