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

Authors :
Dongqing Pan
Ryo Oyama
Tomomi Sato
Takanori Nakane
Ryo Mizunuma
Keita Matsuoka
Yasumasa Joti
Kensuke Tono
Eriko Nango
So Iwata
Toru Nakatsu
Hiroaki Kato
Source :
IUCrJ, Vol 9, Iss 1, Pp 134-145 (2022)
Publication Year :
2022
Publisher :
International Union of Crystallography, 2022.

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 Šis 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.

Details

Language :
English
ISSN :
20522525
Volume :
9
Issue :
1
Database :
Directory of Open Access Journals
Journal :
IUCrJ
Publication Type :
Academic Journal
Accession number :
edsdoj.46bdada9d6934983b448a8f4996580df
Document Type :
article
Full Text :
https://doi.org/10.1107/S2052252521011611