title: Gentamicin B1 is a minor gentamicin component with major nonsense mutation suppression activity.
abstract: Nonsense mutations underlie about 10% of rare genetic disease cases. They introduce a premature termination codon (PTC) and prevent the formation of full-length protein. Pharmaceutical gentamicin, a mixture of several related aminoglycosides, is a frequently used antibiotic in humans that can induce PTC readthrough and suppress nonsense mutations at high concentrations. However, testing of gentamicin in clinical trials has shown that safe doses of this drug produce weak and variable readthrough activity that is insufficient for use as therapy. In this study we show that the major components of pharmaceutical gentamicin lack PTC readthrough activity but the minor component gentamicin B1 (B1) is a potent readthrough inducer. Molecular dynamics simulations reveal the importance of ring I of B1 in establishing a ribosome configuration that permits pairing of a near-cognate complex at a PTC. B1 induced readthrough at all three nonsense codons in cultured cancer cells withTP53(tumor protein p53) mutations, in cells from patients with nonsense mutations in theTPP1(tripeptidyl peptidase 1),DMD(dystrophin),SMARCAL1(SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily a-like 1), andCOL7A1(collagen type VII alpha 1 chain) genes, and in an in vivo tumor xenograft model. The B1 content of pharmaceutical gentamicin is highly variable and major gentamicins suppress the PTC readthrough activity of B1. Purified B1 provides a consistent and effective source of PTC readthrough activity to study the potential of nonsense suppression for treatment of rare genetic disorders.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC, Canada V6T 1Z3.
authors.forename: Alireza
authors.initials: A
authors.lastname: Baradaran-Heravi
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC, Canada V6T 1Z3.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Niesser
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC, Canada V6T 1Z3.
authors.forename: Aruna D
authors.initials: AD
authors.lastname: Balgi
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC, Canada V6T 1Z3.
authors.forename: Kunho
authors.initials: K
authors.lastname: Choi
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC, Canada V6T 1Z3.
authors.forename: Carla
authors.initials: C
authors.lastname: Zimmerman
authors.affiliation: Department of Dermatology and Cutaneous Biology, Thomas Jefferson University, Philadelphia, PA 19107.
authors.forename: Andrew P
authors.initials: AP
authors.lastname: South
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC, Canada V6T 1Z3.
authors.forename: Hilary J
authors.initials: HJ
authors.lastname: Anderson
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC, Canada V6T 1Z3.
authors.forename: Natalie C
authors.initials: NC
authors.lastname: Strynadka
authors.affiliation: Department of Experimental Therapeutics, British Columbia Cancer Agency, Vancouver, BC, Canada V5Z 1L3.
authors.forename: Marcel B
authors.initials: MB
authors.lastname: Bally
authors.affiliation: Department of Biochemistry and Molecular Biology, University of British Columbia, Vancouver, BC, Canada V6T 1Z3; michelr@mail.ubc.ca.
authors.forename: Michel
authors.initials: M
authors.lastname: Roberge
mesh_terms: D000626:Aminopeptidases / Q000235:genetics; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000494:pharmacology*; D045744:Cell Line, Tumor; D018389:Codon, Nonsense / Q000235:genetics*; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000235:genetics; D016189:Dystrophin / Q000235:genetics; D005839:Gentamicins / Q000737:chemistry / Q000494:pharmacology*; D006801:Humans; D009154:Mutation / Q000187:drug effects*; D057057:Serine Proteases / Q000235:genetics; D000091346:Tripeptidyl-Peptidase 1; D016159:Tumor Suppressor Protein p53 / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D018389:Codon, Nonsense; C484258:DMD protein, human; D016189:Dystrophin; D005839:Gentamicins; D000091346:Tripeptidyl-Peptidase 1; D016159:Tumor Suppressor Protein p53; C023804:gentamicin B; D057057:Serine Proteases; D000626:Aminopeptidases; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; C000717267:TPP1 protein, human
keywords: cancer; gentamicin B1; nonsense mutation; premature stop codon readthrough; rare genetic diseases