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Aminoglycoside ribosome interactions reveal novel conformational states at ambient temperature

Mary E. O’Sullivan, Frédéric Poitevin, Raymond G. Sierra, Cornelius Gati, E. Han Dao, Yashas Rao, Fulya Aksit, Halilibrahim Ciftci, Nicholas Corsepius, Robert Greenhouse, Brandon Hayes, Mark S. Hunter, Mengling Liang, Alex McGurk, Paul Mbgam, Trevor Obrinsky, Fátima Pardo-Avila, Matt Seaberg, Alan G. Cheng, Anthony J. Ricci, Hasan DeMirci
doi: https://doi.org/10.1101/372144
Mary E. O’Sullivan
Stanford University School of Medicine, Department of Otolaryngology-Head and Neck Surgery, Palo Alto, California, USA
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Frédéric Poitevin
Stanford University, Department of Structural Biology, Palo Alto, California, USAStanford PULSE Institute, SLAC National Laboratory, Menlo Park, California, USA
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Raymond G. Sierra
Linac Coherent Light Source, SLAC National Laboratory, Menlo Park, California, USA
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Cornelius Gati
Stanford University, Department of Structural Biology, Palo Alto, California, USABiosciences Division, SLAC National Laboratory, Menlo Park, California, USA
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E. Han Dao
Stanford PULSE Institute, SLAC National Laboratory, Menlo Park, California, USA
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Yashas Rao
Linac Coherent Light Source, SLAC National Laboratory, Menlo Park, California, USA
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Fulya Aksit
Linac Coherent Light Source, SLAC National Laboratory, Menlo Park, California, USA
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Halilibrahim Ciftci
Linac Coherent Light Source, SLAC National Laboratory, Menlo Park, California, USA
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Nicholas Corsepius
Stanford University, Department of Structural Biology, Palo Alto, California, USA
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Robert Greenhouse
Stanford University School of Medicine, Department of Otolaryngology-Head and Neck Surgery, Palo Alto, California, USA
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Brandon Hayes
Linac Coherent Light Source, SLAC National Laboratory, Menlo Park, California, USA
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Mark S. Hunter
Linac Coherent Light Source, SLAC National Laboratory, Menlo Park, California, USA
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Mengling Liang
Linac Coherent Light Source, SLAC National Laboratory, Menlo Park, California, USA
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Alex McGurk
Linac Coherent Light Source, SLAC National Laboratory, Menlo Park, California, USA
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Paul Mbgam
Linac Coherent Light Source, SLAC National Laboratory, Menlo Park, California, USA
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Trevor Obrinsky
Linac Coherent Light Source, SLAC National Laboratory, Menlo Park, California, USA
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Fátima Pardo-Avila
Stanford University, Department of Structural Biology, Palo Alto, California, USA
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Matt Seaberg
Linac Coherent Light Source, SLAC National Laboratory, Menlo Park, California, USA
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Alan G. Cheng
Stanford University School of Medicine, Department of Otolaryngology-Head and Neck Surgery, Palo Alto, California, USA
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Anthony J. Ricci
Stanford University School of Medicine, Department of Otolaryngology-Head and Neck Surgery, Palo Alto, California, USA
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  • For correspondence: Hasan_DeMirci@Stanford.edu aricci@Stanford.edu
Hasan DeMirci
Stanford University, Department of Structural Biology, Palo Alto, California, USAStanford PULSE Institute, SLAC National Laboratory, Menlo Park, California, USABiosciences Division, SLAC National Laboratory, Menlo Park, California, USA
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  • For correspondence: Hasan_DeMirci@Stanford.edu aricci@Stanford.edu
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ABSTRACT

The bacterial 30S ribosomal subunit is a primary antibiotic target. Despite decades of discovery, the mechanisms by which antibiotic binding induces ribosomal dysfunction are not fully understood. Ambient temperature crystallographic techniques allow more biologically relevant investigation of how local antibiotic binding site interactions trigger global subunit rearrangements that perturb protein synthesis. Here, the structural effects of 2-deoxystreptamine (paromomycin and sisomicin), a novel sisomicin derivative, N1-methyl sulfonyl sisomicin (N1MS) and the non-deoxystreptamine (streptomycin) aminoglycosides on the ribosome at ambient and cryogenic temperatures were examined. Comparative studies led to three main observations. First, individual aminoglycoside-ribosome interactions in the decoding center were similar for cryogenic vs ambient temperature structures. Second, analysis of a highly conserved GGAA tetraloop of h45 revealed aminoglycoside-specific conformational changes, which are affected by temperature only for N1MS. We report the h44/h45 interface in varying states, that is, engaged, disengaged and in equilibrium. Thirdly, we observe aminoglycoside-induced effects on 30S domain closure, including a novel intermediary closure state, which is also sensitive to temperature. Analysis of three ambient and five cryogenic crystallography datasets reveal a correlation between h44/h45 engagement and domain closure. These observations illustrate the role of ambient temperature crystallography in identifying dynamic mechanisms of ribosomal dysfunction induced by local drug-binding site interactions. Together these data identify tertiary ribosomal structural changes induced by aminoglycoside binding that provides functional insight and targets for drug design.

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Posted July 19, 2018.
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Aminoglycoside ribosome interactions reveal novel conformational states at ambient temperature
Mary E. O’Sullivan, Frédéric Poitevin, Raymond G. Sierra, Cornelius Gati, E. Han Dao, Yashas Rao, Fulya Aksit, Halilibrahim Ciftci, Nicholas Corsepius, Robert Greenhouse, Brandon Hayes, Mark S. Hunter, Mengling Liang, Alex McGurk, Paul Mbgam, Trevor Obrinsky, Fátima Pardo-Avila, Matt Seaberg, Alan G. Cheng, Anthony J. Ricci, Hasan DeMirci
bioRxiv 372144; doi: https://doi.org/10.1101/372144
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Aminoglycoside ribosome interactions reveal novel conformational states at ambient temperature
Mary E. O’Sullivan, Frédéric Poitevin, Raymond G. Sierra, Cornelius Gati, E. Han Dao, Yashas Rao, Fulya Aksit, Halilibrahim Ciftci, Nicholas Corsepius, Robert Greenhouse, Brandon Hayes, Mark S. Hunter, Mengling Liang, Alex McGurk, Paul Mbgam, Trevor Obrinsky, Fátima Pardo-Avila, Matt Seaberg, Alan G. Cheng, Anthony J. Ricci, Hasan DeMirci
bioRxiv 372144; doi: https://doi.org/10.1101/372144

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