Biopolym. Cell. 2023; 39(4):265-276.
Structure and Function of Biopolymers
Structural characterization of cephaeline binding to the eukaryotic ribosome using Cryo-Electron Microscopy
1Kolosova O., 1Zgadzay Y., 2Stetsenko A., 2Atamas A., 3Wu C., 3Sachs M. S., 1Jenner L., 2Guskov A., 1Yusupov M.
- Department of Integrated Structural Biology, Institute of Genetics and Molecular and Cellular Biology
University of Strasbourg, Illkirch, 67400, France - Groningen Biomolecular Sciences and Biotechnology Institute (GBB), University of Groningen
Groningen, 9747 AG, the Netherlands - Department of Biology, Texas A&M University
College Station, TX, USA
Abstract
Aim. In our work, we aimed to elucidate the binding mechanism of another alkaloid, cephaeline, to the eukaryotic ribosome. Methods. We used cryogenic electron microscopy and cell-free assays to reveal its mechanism of action. Results. Our results indicate that cephaeline binds to the E-tRNA binding site on the small subunit of the eukaryotic ribosome. Similar to emetine, cephaeline forms a stacking interaction with G841 of 18S rRNA and L132 of the protein uS11. We propose the hypothesis of cephaeline specificity to eukaryotes by comparing the interaction pattern of cephaeline with other inhibitors binding to the E-site of the mRNA tunnel. Conclusions. The high-resolution structure of ribosome-bound cephaeline (2.45 Å) allowed us to precisely determine the inhibitor’s position in the binding site, which holds potential for the development of the next generation of drugs targeting the mRNA tunnel of the ribosome.
Keywords: ribosome, cephaeline, anti-protozoan, cryo-EM
Full text: (PDF, in English)
References
[1]
Lin J, Zhou D, Steitz TA, Polikanov YS, Gagnon MG. Ribosome-Targeting Antibiotics: Modes of Action, Mechanisms of Resistance, and Implications for Drug Design. Annu Rev Biochem. 2018; 87:451-78.
[2]
Dmitriev SE, Vladimirov DO, Lashkevich KA. A Quick Guide to Small-Molecule Inhibitors of Eukaryotic Protein Synthesis. Biochemistry (Mosc). 2020; 85(11):1389-421.
[3]
Watson ZL, Ward FR, Méheust R, Ad O, Schepartz A, Banfield JF, Cate JH. Structure of the bacterial ribosome at 2 Å resolution. Elife. 2020; 9:e60482.
[4]
Fromm SA, O'Connor KM, Purdy M, Bhatt PR, Loughran G, Atkins JF, Jomaa A, Mattei S. The translating bacterial ribosome at 1.55 Å resolution generated by cryo-EM imaging services. Nat Commun. 2023; 14(1):1095.
[5]
Xue L, Lenz S, Zimmermann-Kogadeeva M, Tegunov D, Cramer P, Bork P, Rappsilber J, Mahamid J. Visualizing translation dynamics at atomic detail inside a bacterial cell. Nature. 2022; 610(7930):205-11.
[6]
Xing H, Taniguchi R, Khusainov I, Kreysing JP, Welsch S, Turoňová B, Beck M. Translation dynamics in human cells visualized at high resolution reveal cancer drug action. Science. 2023; 381(6653):70-5.
[7]
Ben-Shem A, Garreau de Loubresse N, Melnikov S, Jenner L, Yusupova G, Yusupov M. The structure of the eukaryotic ribosome at 3.0 Å resolution. Science. 2011; 334(6062):1524-9.
[8]
Bansal D, Sehgal R, Chawla Y, Mahajan RC, Malla N. In vitro activity of antiamoebic drugs against clinical isolates of Entamoeba histolytica and Entamoeba dispar. Ann Clin Microbiol Antimicrob. 2004; 3:27.
[9]
Matthews H, Usman-Idris M, Khan F, Read M, Nirmalan N. Drug repositioning as a route to anti-malarial drug discovery: preliminary investigation of the in vitro anti-malarial efficacy of emetine dihydrochloride hydrate. Malar J. 2013; 12:359.
[10]
Zgadzay Y, Kolosova O, Stetsenko A, Wu C, Bruchlen D, Usachev K, Validov S, Jenner L, Rogachev A, Yusupova G, Sachs MS, Guskov A, Yusupov M. E-site drug specificity of the human pathogen Candida albicans ribosome. Sci Adv. 2022; 8(21):eabn1062.
[11]
Lee MR. Ipecacuanha: the South American vomiting root. J R Coll Physicians Edinb. 2008; 38(4):355-60.
[12]
Lemmens-Gruber R, Karkhaneh A, Studenik C, Heistracher P. Cardiotoxicity of emetine dihydrochloride by calcium channel blockade in isolated preparations and ventricular myocytes of guinea-pig hearts. Br J Pharmacol. 1996; 117(2):377-83.
[13]
Panwar P, Burusco KK, Abubaker M, Matthews H, Gutnov A, Fernández-Álvaro E, Bryce RA, Wilkinson J, Nirmalan N. Lead Optimization of Dehydroemetine for Repositioned Use in Malaria. Antimicrob Agents Chemother. 2020; 64(4):e01444-19.
[14]
Maddison JE, Page SW, Church DB. Small animal clinical pharmacology.
[15]
Ren PX, Shang WJ, Yin WC, Ge H, Wang L, Zhang XL, Li BQ, Li HL, Xu YC, Xu EH, Jiang HL, Zhu LL, Zhang LK, Bai F. A multi-targeting drug design strategy for identifying potent anti-SARS-CoV-2 inhibitors. Acta Pharmacol Sin. 2022; 43(2):483-93.
[16]
Gupta RS, Siminovitch L. Mutants of CHO cells resistant to the protein synthesis inhibitors, cryptopleurine and tylocrebrine: genetic and biochemical evidence for common site of action of emetine, cryptopleurine, tylocrebine, and tubulosine. Biochemistry. 1977; 16(14):3209-14.
[17]
Chang S, Wasmuth JJ. Construction and characterization of Chinese hamster cell EmtA EmtB double mutants. Mol Cell Biol. 1983; 3(5):761-72.
[18]
Chang S, Wasmuth JJ. Genetic and biochemical distinction among Chinese hamster cell emtA, emtB, and emtC mutants. Mol Cell Biol. 1983; 3(3):429-38.
[19]
Rhoads DD, Roufa DJ. Emetine resistance of Chinese hamster cells: structures of wild-type and mutant ribosomal protein S14 mRNAs. Mol Cell Biol. 1985; 5(7):1655-9.
[20]
Grant P, Sánchez L, Jiménez A. Cryptopleurine resistance: genetic locus for a 40S ribosomal component in Saccharomyces cerevisiae. J Bacteriol. 1974; 120(3):1308-14.
[21]
Madjar JJ, Nielsen-Smith K, Frahm M, Roufa DJ. Emetine resistance in chinese hamster ovary cells is associated with an altered ribosomal protein S14 mRNA. Proc Natl Acad Sci U S A. 1982; 79(4):1003-7.
[22]
Wong W, Bai XC, Brown A, Fernandez IS, Hanssen E, Condron M, Tan YH, Baum J, Scheres SH. Cryo-EM structure of the Plasmodium falciparum 80S ribosome bound to the anti-protozoan drug emetine. Elife. 2014; 3:e03080.
[23]
Bucher K, Skogerson L. Cryptopleurine--an inhibitor of translocation. Biochemistry. 1976; 15(22):4755-9.
[24]
Brodersen DE, Clemons WM Jr, Carter AP, Morgan-Warren RJ, Wimberly BT, Ramakrishnan V. The structural basis for the action of the antibiotics tetracycline, pactamycin, and hygromycin B on the 30S ribosomal subunit. Cell. 2000; 103(7):1143-54.
[25]
Garreau de Loubresse N, Prokhorova I, Holtkamp W, Rodnina MV, Yusupova G, Yusupov M. Structural basis for the inhibition of the eukaryotic ribosome. Nature. 2014; 513(7519):517-22.
[26]
Polikanov YS, Osterman IA, Szal T, Tashlitsky VN, Serebryakova MV, Kusochek P, Bulkley D, Malanicheva IA, Efimenko TA, Efremenkova OV, Konevega AL, Shaw KJ, Bogdanov AA, Rodnina MV, Dontsova OA, Mankin AS, Steitz TA, Sergiev PV. Amicoumacin a inhibits translation by stabilizing mRNA interaction with the ribosome. Mol Cell. 2014; 56(4):531-40.
[27]
Prokhorova IV, Akulich KA, Makeeva DS, Osterman IA, Skvortsov DA, Sergiev PV, Dontsova OA, Yusupova G, Yusupov MM, Dmitriev SE. Amicoumacin A induces cancer cell death by targeting the eukaryotic ribosome. Sci Rep. 2016; 6:27720.
[28]
Li X, Zheng S, Agard DA, Cheng Y. Asynchronous data acquisition and on-the-fly analysis of dose fractionated cryoEM images by UCSFImage. J Struct Biol. 2015; 192(2):174-8.
[29]
Punjani A, Rubinstein JL, Fleet DJ, Brubaker MA. cryoSPARC: algorithms for rapid unsupervised cryo-EM structure determination. Nat Methods. 2017; 14(3):290-6.
[30]
Liebschner D, Afonine PV, Baker ML, Bunkóczi G, Chen VB, Croll TI, Hintze B, Hung LW, Jain S, McCoy AJ, Moriarty NW, Oeffner RD, Poon BK, Prisant MG, Read RJ, Richardson JS, Richardson DC, Sammito MD, Sobolev OV, Stockwell DH, Terwilliger TC, Urzhumtsev AG, Videau LL, Williams CJ, Adams PD. Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix. Acta Crystallogr D Struct Biol. 2019; 75(Pt 10):861-77.
[31]
Emsley P, Lohkamp B, Scott WG, Cowtan K. Features and development of Coot. Acta Crystallogr D Biol Crystallogr. 2010; 66(Pt 4):486-501.
[32]
Goddard TD, Huang CC, Meng EC, Pettersen EF, Couch GS, Morris JH, Ferrin TE. UCSF ChimeraX: Meeting modern challenges in visualization and analysis. Protein Sci. 2018; 27(1):14-25.
[33]
Pettersen EF, Goddard TD, Huang CC, Meng EC, Couch GS, Croll TI, Morris JH, Ferrin TE. UCSF ChimeraX: Structure visualization for researchers, educators, and developers. Protein Sci. 2021; 30(1):70-82.
