Crystallographic characterization of the ribosomal binding site and molecular mechanism of action of Hygromycin A

Kaminishi, T.; Schedlbauer, Andreas; Fabbretti, Attilio; Brandi, Letizia; Ochoa-Lizarralde, B.; He, Cheng-Guang; Milón, Pohl; Connell, Sean R.; Gualerzi, Claudio O.; Fucini, Paola

doi:10.1093/nar/gkv975

Cited by 14 publications

(19 citation statements)

References 65 publications

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“…In addition, our IF3 DL -30S reporter assay can provide novel aspects of the inhibiting mechanism of known 30S-binding drugs. Similar approaches have allowed detailed descriptions for other inhibitors of the ribosome [54,55]. …”

Section: Discussionmentioning

confidence: 99%

Conformational Response of 30S-bound IF3 to A-Site Binders Streptomycin and Kanamycin

et al. 2016

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Aminoglycoside antibiotics are widely used to treat infectious diseases. Among them, streptomycin and kanamycin (and derivatives) are of importance to battle multidrug-resistant (MDR) Mycobacterium tuberculosis. Both drugs bind the small ribosomal subunit (30S) and inhibit protein synthesis. Genetic, structural, and biochemical studies indicate that local and long-range conformational rearrangements of the 30S subunit account for this inhibition. Here, we use intramolecular FRET between the C- and N-terminus domains of the flexible IF3 to monitor real-time perturbations of their binding sites on the 30S platform. Steady and pre-steady state binding experiments show that both aminoglycosides bring IF3 domains apart, promoting an elongated state of the factor. Binding of Initiation Factor IF1 triggers closure of IF3 bound to the 30S complex, while both aminoglycosides revert the IF1-dependent conformation. Our results uncover dynamic perturbations across the 30S subunit, from the A-site to the platform, and suggest that both aminoglycosides could interfere with prokaryotic translation initiation by modulating the interaction between IF3 domains with the 30S platform.

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Section: Discussionmentioning

confidence: 99%

Conformational Response of 30S-bound IF3 to A-Site Binders Streptomycin and Kanamycin

et al. 2016

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“…PASS online predicted molecule 1 as a small ribosomal subunit inhibitor, whereas molecule 1 binds to large ribosomal subunit and inhibits its peptidyl transferase activity (Kaminishi et al, 2015).…”

Section: Resultsmentioning

confidence: 99%

Reliable Target Prediction of Bioactive Molecules Based on Chemical Similarity Without Employing Statistical Methods

et al. 2019

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The prediction of biological targets of bioactive molecules from machine-readable materials can be routinely performed by computational target prediction tools (CTPTs). However, the prediction of biological targets of bioactive molecules from non-digital materials (e.g., printed or handwritten documents) has not been possible due to the complex nature of bioactive molecules and impossibility of employing computations. Improving the target prediction accuracy is the most important challenge for computational target prediction. A minimum structure is identified for each group of neighbor molecules in the proposed method. Each group of neighbor molecules represents a distinct structural class of molecules with the same function in relation to the target. The minimum structure is employed as a query to search for molecules that perfectly satisfy the minimum structure of what is guessed crucial for the targeted activity. The proposed method is based on chemical similarity, but only molecules that perfectly satisfy the minimum structure are considered. Structurally related bioactive molecules found with the same minimum structure were considered as neighbor molecules of the query molecule. The known target of the neighbor molecule is used as a reference for predicting the target of the neighbor molecule with an unknown target. A lot of information is needed to identify the minimum structure, because it is necessary to know which part(s) of the bioactive molecule determines the precise target or targets responsible for the observed phenotype. Therefore, the predicted target based on the minimum structure without employing the statistical significance is considered as a reliable prediction. Since only molecules that perfectly (and not partly) satisfy the minimum structure are considered, the minimum structure can be used without similarity calculations in non-digital materials and with similarity calculations (perfect similarity) in machine-readable materials. Nine tools (PASS online, PPB, SEA, TargetHunter, PharmMapper, ChemProt, HitPick, SuperPred, and SPiDER), which can be used for computational target prediction, are compared with the proposed method for 550 target predictions. The proposed method, SEA, PPB, and PASS online, showed the best quality and quantity for the accurate predictions.

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“…Until recently, it was thought that most of the ribosome-targeting drugs inhibit protein synthesis by altering the conformation of the ribosome or by preventing the proper binding of the ribosomal substrates. This paradigm, however, was challenged by recent structural studies of antibiotics blasticidin S ( 38 ), amicoumacin A ( 20 ), negamycin ( 39 , 40 ) and hygromycin A ( 41 , 42 ). These antibiotics were shown to tether the ligands (such as mRNA and tRNAs) to the ribosome or force their non-productive conformations.…”

Section: Discussionmentioning

confidence: 99%

Insights into RNA binding by the anticancer drug cisplatin from the crystal structure of cisplatin-modified ribosome

et al. 2016

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Cisplatin is a widely prescribed anticancer drug, which triggers cell death by covalent binding to a broad range of biological molecules. Among cisplatin targets, cellular RNAs remain the most poorly characterized molecules. Although cisplatin was shown to inactivate essential RNAs, including ribosomal, spliceosomal and telomeric RNAs, cisplatin binding sites in most RNA molecules are unknown, and therefore it remains challenging to study how modifications of RNA by cisplatin contributes to its toxicity. Here we report a 2.6Å-resolution X-ray structure of cisplatin-modified 70S ribosome, which describes cisplatin binding to the ribosome and provides the first nearly atomic model of cisplatin–RNA complex. We observe nine cisplatin molecules bound to the ribosome and reveal consensus structural features of the cisplatin-binding sites. Two of the cisplatin molecules modify conserved functional centers of the ribosome—the mRNA-channel and the GTPase center. In the mRNA-channel, cisplatin intercalates between the ribosome and the messenger RNA, suggesting that the observed inhibition of protein synthesis by cisplatin is caused by impaired mRNA-translocation. Our structure provides an insight into RNA targeting and inhibition by cisplatin, which can help predict cisplatin-binding sites in other cellular RNAs and design studies to elucidate a link between RNA modifications by cisplatin and cisplatin toxicity.

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Crystallographic characterization of the ribosomal binding site and molecular mechanism of action of Hygromycin A

Cited by 14 publications

References 65 publications

Conformational Response of 30S-bound IF3 to A-Site Binders Streptomycin and Kanamycin

Conformational Response of 30S-bound IF3 to A-Site Binders Streptomycin and Kanamycin

Reliable Target Prediction of Bioactive Molecules Based on Chemical Similarity Without Employing Statistical Methods

Insights into RNA binding by the anticancer drug cisplatin from the crystal structure of cisplatin-modified ribosome

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