Advanced Photon Source

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How Anticancer Drugs Enter Cells

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Nucleosides are precursors of nucleic acids, are involved in many biochemical processes in cells, especially in the storage and transfer of genetic information. Concentrative nucleoside transporters (CNTs) play an important role in transporting nucleosides and nucleoside-derived drugs into cells. Although scientific advances have recognized the elevator model as the mechanism of this transport process, scientists have only identified the structures of the two end states of the transporter that exist at the start and end of the transport cycle; the more detailed structural changes involved throughout the entire cycle have remained somewhat of a mystery. Researchers set out to further investigate the transport cycle utilizing data collected at the U.S. Department of Energy’s Advanced Photon Source (APS) and characterized most of the structural changes, including novel intermediate-state conformations, that occur in the CNT throughout its transport cycle as it moves nucleosides and nucleoside analogs across the cell membrane. The results of this study could help scientists better understand the elevator model for transporting various molecules into cells, and could also help with discovery of new nucleoside-derived drugs such as anticancer and antiviral therapies.

Nucleoside analogs represent an important class of compounds that are used clinically, as anticancer drugs and antiviral drugs, for example. These analogs are essentially modified nucleosides that have antitumor or antiviral properties because of their ability to block DNA synthesis. The CNT proteins are used to help nucleosides and their analogs enter cells. These transporters use an ion gradient as an energy source to transport nucleosides and nucleoside-derived drugs against their chemical gradients across cell membranes into cells.

The elevator model represents an emerging mechanism of the transport process, in which a region of the CNT known as the substrate-binding transport domain moves a large distance across the membrane. This mechanism has been characterized by a transition between two states, but the conformational path that leads to the transition has remained unknown. This is mostly because the available structural information has been limited to the two end states of the CNT that exist at the start and end of the transport cycle.

Using data collected at the Southeast Regional Collaborative Access Team 22-ID-D beamline and the Northeastern Collaborative Access Team 24-ID-C beamline, both at the APS, the researchers from Duke University Medical Center captured and visualized the movements of the CNT in a time-lapse manner, helping them to better understand how this transporter works. The APS is an Office of Science user facility at Argonne National Laboratory.

Although they had previously tried to capture alternate conformations of the CNT during transport, most of their approaches had failed. This study, however, was the first to provide a visualization of almost all stages of the elevator model. They determined the structures of nearly all the shapes of the CNT in motion, providing a trajectory of its conformational transitions in the elevator model (Fig. 1). These findings showed that multiple intermediate steps and state-dependent conformational changes occur within the transport domain as the CNT slowly moves its cargo like an elevator, stopping at different points across the cell membrane before reaching the inside of the cell.

The researchers were initially surprised when they identified these intermediate steps. It had traditionally been believed that the transition between the starting and ending states of the CNT was transient, and that the transport cycle did not involve intermediate states. However, the more the researchers analyzed the novel intermediate state structures, the more they realized that these intermediate conformational changes do occur and play important roles in the transport cycle. Their subsequent biochemical studies are consistent with their structural observation of the importance of the intermediate states. They now hope to capture a few more of these intermediate conformations to help map the entire conformational landscape of the transport cycle used by this transporter.

This more detailed understanding of the elevator model can help guide development of new anticancer and antiviral drugs that are more selective and more efficient.  — Nicola Parry

See: Marscha Hirschi, Zachary Lee Johnson, and Seok-Yong Lee*, “Visualizing multistep elevator-like transitions of a nucleoside transporter,” Nature 545, 66 (4 May 2017). DOI: 10.1038/nature22057

Author affiliation: Duke University Medical Center

Correspondence: *seok-yong.lee@duke.edu

This work was supported by National Institutes of Health (NIH) R01 GM100984 (S.-Y.L.) and NIH R35 NS097241 (S.-Y.L.). Supporting institutions for the Southeast Regional Collaborative Access Team may be found at www.ser-cat.org/members.html. Northeastern Collaborative Access Team beamlines, which are funded by the National Institute of General Medical Sciences from the National Institutes of Health (P41 GM103403). The Pilatus 6M detector on 24-ID-C beam line is funded by a NIH-ORIP HEI grant (S10 RR029205). This research used resources of the Advanced Photon Source, a U.S. Department of Energy (DOE) Office of Science User Facility operated for the DOE Office of Science by Argonne National Laboratory under Contract No. DE-AC02-06CH11357.

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Published Date: 
02.02.2018