Chemically related 4,5-linked aminoglycoside antibiotics drive subunit rotation in opposite directions

Michael R. Wasserman; Arto Pulk; Zhou Zhou; Roger B. Altman; John C. Zinder; Keith D. Green; Sylvie Garneau-Tsodikova; Jamie H. Doudna Cate; Scott C. Blanchard

doi:10.1038/ncomms8896

Chemically related 4,5-linked aminoglycoside antibiotics drive subunit rotation in opposite directions

Michael R. Wasserman, Arto Pulk, Zhou Zhou, Roger B. Altman, John C. Zinder, Keith D. Green, Sylvie Garneau-Tsodikova, Jamie H. Doudna Cate, Scott C. Blanchard

Research output: Contribution to journal › Article › peer-review

52 Scopus citations

Abstract

Dynamic remodelling of intersubunit bridge B2, a conserved RNA domain of the bacterial ribosome connecting helices 44 (h44) and 69 (H69) of the small and large subunit, respectively, impacts translation by controlling intersubunit rotation. Here we show that aminoglycosides chemically related to neomycin - paromomycin, ribostamycin and neamine - each bind to sites within h44 and H69 to perturb bridge B2 and affect subunit rotation. Neomycin and paromomycin, which only differ by their ring-I 6′-polar group, drive subunit rotation in opposite directions. This suggests that their distinct actions hinge on the 6′-substituent and the drugâ €™ s net positive charge. By solving the crystal structure of the paromomycin-ribosome complex, we observe specific contacts between the apical tip of H69 and the 6′-hydroxyl on paromomycin from within the drugâ €™ s canonical h44-binding site. These results indicate that aminoglycoside actions must be framed in the context of bridge B2 and their regulation of subunit rotation.

Original language	English
Article number	7896
Journal	Nature Communications
Volume	6
DOIs	https://doi.org/10.1038/ncomms8896
State	Published - Jul 30 2015

Bibliographical note

Funding Information:
We thank R. Green (Johns Hopkins University) for providing the S13 knockout strain, P. Schultz for providing the L5 knockout strain (Scripps Research Institute), T. Suzuki (University of Tokyo) for providing the pKK3535 ribosome plasmids, D. Crish (Wayne State University) for providing 6′-deoxy-paromomycin and M. O’Connor (University of Missouri-Kansas City) for helpful discussions throughout the course of this work. We also acknowledge helpful discussions and insights provided by all members of the Blanchard and Cate laboratories and J. Headd (Lawrence Berkeley National Laboratory) for help with PHENIX refinement. This work was supported by the US National Institutes of Health (2R01GM079238 to S.C.B., 1R01GM65050 to J.H.D.C., 1R01AI090048 to S.G.-T. and National Cancer Institute grant CA92584 for the SIBYLS and 8.3.1 beamlines at the Advanced Light Source (ALS), Lawrence Berkeley National Laboratory), the Human Frontiers in Science Program (RGY0088), the National Science Foundation (0644129) and the US Department of Energy (DE-AC0376SF00098 for the SIBYLS and 8.3.1 beamlines at the ALS, Lawrence Berkeley National Laboratory).

ASJC Scopus subject areas

General Chemistry
General Biochemistry, Genetics and Molecular Biology
General Physics and Astronomy

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title = "Chemically related 4,5-linked aminoglycoside antibiotics drive subunit rotation in opposite directions",

abstract = "Dynamic remodelling of intersubunit bridge B2, a conserved RNA domain of the bacterial ribosome connecting helices 44 (h44) and 69 (H69) of the small and large subunit, respectively, impacts translation by controlling intersubunit rotation. Here we show that aminoglycosides chemically related to neomycin - paromomycin, ribostamycin and neamine - each bind to sites within h44 and H69 to perturb bridge B2 and affect subunit rotation. Neomycin and paromomycin, which only differ by their ring-I 6′-polar group, drive subunit rotation in opposite directions. This suggests that their distinct actions hinge on the 6′-substituent and the drug{\^a} €{\texttrademark} s net positive charge. By solving the crystal structure of the paromomycin-ribosome complex, we observe specific contacts between the apical tip of H69 and the 6′-hydroxyl on paromomycin from within the drug{\^a} €{\texttrademark} s canonical h44-binding site. These results indicate that aminoglycoside actions must be framed in the context of bridge B2 and their regulation of subunit rotation.",

author = "Wasserman, {Michael R.} and Arto Pulk and Zhou Zhou and Altman, {Roger B.} and Zinder, {John C.} and Green, {Keith D.} and Sylvie Garneau-Tsodikova and {Doudna Cate}, {Jamie H.} and Blanchard, {Scott C.}",

note = "Funding Information: We thank R. Green (Johns Hopkins University) for providing the S13 knockout strain, P. Schultz for providing the L5 knockout strain (Scripps Research Institute), T. Suzuki (University of Tokyo) for providing the pKK3535 ribosome plasmids, D. Crish (Wayne State University) for providing 6′-deoxy-paromomycin and M. O{\textquoteright}Connor (University of Missouri-Kansas City) for helpful discussions throughout the course of this work. We also acknowledge helpful discussions and insights provided by all members of the Blanchard and Cate laboratories and J. Headd (Lawrence Berkeley National Laboratory) for help with PHENIX refinement. This work was supported by the US National Institutes of Health (2R01GM079238 to S.C.B., 1R01GM65050 to J.H.D.C., 1R01AI090048 to S.G.-T. and National Cancer Institute grant CA92584 for the SIBYLS and 8.3.1 beamlines at the Advanced Light Source (ALS), Lawrence Berkeley National Laboratory), the Human Frontiers in Science Program (RGY0088), the National Science Foundation (0644129) and the US Department of Energy (DE-AC0376SF00098 for the SIBYLS and 8.3.1 beamlines at the ALS, Lawrence Berkeley National Laboratory).",

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AU - Wasserman, Michael R.

AU - Pulk, Arto

AU - Zhou, Zhou

AU - Altman, Roger B.

AU - Zinder, John C.

AU - Green, Keith D.

AU - Garneau-Tsodikova, Sylvie

AU - Doudna Cate, Jamie H.

AU - Blanchard, Scott C.

N1 - Funding Information: We thank R. Green (Johns Hopkins University) for providing the S13 knockout strain, P. Schultz for providing the L5 knockout strain (Scripps Research Institute), T. Suzuki (University of Tokyo) for providing the pKK3535 ribosome plasmids, D. Crish (Wayne State University) for providing 6′-deoxy-paromomycin and M. O’Connor (University of Missouri-Kansas City) for helpful discussions throughout the course of this work. We also acknowledge helpful discussions and insights provided by all members of the Blanchard and Cate laboratories and J. Headd (Lawrence Berkeley National Laboratory) for help with PHENIX refinement. This work was supported by the US National Institutes of Health (2R01GM079238 to S.C.B., 1R01GM65050 to J.H.D.C., 1R01AI090048 to S.G.-T. and National Cancer Institute grant CA92584 for the SIBYLS and 8.3.1 beamlines at the Advanced Light Source (ALS), Lawrence Berkeley National Laboratory), the Human Frontiers in Science Program (RGY0088), the National Science Foundation (0644129) and the US Department of Energy (DE-AC0376SF00098 for the SIBYLS and 8.3.1 beamlines at the ALS, Lawrence Berkeley National Laboratory).

PY - 2015/7/30

Y1 - 2015/7/30

N2 - Dynamic remodelling of intersubunit bridge B2, a conserved RNA domain of the bacterial ribosome connecting helices 44 (h44) and 69 (H69) of the small and large subunit, respectively, impacts translation by controlling intersubunit rotation. Here we show that aminoglycosides chemically related to neomycin - paromomycin, ribostamycin and neamine - each bind to sites within h44 and H69 to perturb bridge B2 and affect subunit rotation. Neomycin and paromomycin, which only differ by their ring-I 6′-polar group, drive subunit rotation in opposite directions. This suggests that their distinct actions hinge on the 6′-substituent and the drugâ €™ s net positive charge. By solving the crystal structure of the paromomycin-ribosome complex, we observe specific contacts between the apical tip of H69 and the 6′-hydroxyl on paromomycin from within the drugâ €™ s canonical h44-binding site. These results indicate that aminoglycoside actions must be framed in the context of bridge B2 and their regulation of subunit rotation.

AB - Dynamic remodelling of intersubunit bridge B2, a conserved RNA domain of the bacterial ribosome connecting helices 44 (h44) and 69 (H69) of the small and large subunit, respectively, impacts translation by controlling intersubunit rotation. Here we show that aminoglycosides chemically related to neomycin - paromomycin, ribostamycin and neamine - each bind to sites within h44 and H69 to perturb bridge B2 and affect subunit rotation. Neomycin and paromomycin, which only differ by their ring-I 6′-polar group, drive subunit rotation in opposite directions. This suggests that their distinct actions hinge on the 6′-substituent and the drugâ €™ s net positive charge. By solving the crystal structure of the paromomycin-ribosome complex, we observe specific contacts between the apical tip of H69 and the 6′-hydroxyl on paromomycin from within the drugâ €™ s canonical h44-binding site. These results indicate that aminoglycoside actions must be framed in the context of bridge B2 and their regulation of subunit rotation.

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Chemically related 4,5-linked aminoglycoside antibiotics drive subunit rotation in opposite directions

Abstract

Bibliographical note

ASJC Scopus subject areas

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