Viomycin

Member of the tuberactinomycin family of antibiotics. Inhibits group I intron splicing and prokaryotic protein synthesis. Freezes bacterial ribosomes in either the pre- or post-translational state. Facilitates trans-cleavage of the Neurospora VS ribozyme.

Molecular basis for the selectivity of antituberculosis compounds capreomycin and viomycin.[Pubmed:21768509]

Antimicrob Agents Chemother. 2011 Oct;55(10):4712-7.

Capreomycin and the structurally similar compound Viomycin are cyclic peptide antibiotics which are particularly active against Mycobacterium tuberculosis, including multidrug resistant strains. Both antibiotics bind across the ribosomal interface involving 23S rRNA helix 69 (H69) and 16S rRNA helix 44 (h44). The binding site of tuberactinomycins in h44 partially overlaps with that of aminoglycosides, and they share with these drugs the side effect of irreversible hearing loss. Here we studied the drug target interaction on ribosomes modified by site-directed mutagenesis. We identified rRNA residues in h44 as the main determinants of phylogenetic selectivity, predict compensatory evolution to impact future resistance development, and propose mechanisms involved in tuberactinomycin ototoxicity, which may enable the development of improved, less-toxic derivatives.

Crystallization and preliminary crystallographic analysis of the first condensation domain of viomycin synthetase.[Pubmed:23545648]

Acta Crystallogr Sect F Struct Biol Cryst Commun. 2013 Apr 1;69(Pt 4):412-5.

Nonribosomal peptide synthetases (NRPSs) are large multimodular enzymes that synthesize important secondary metabolites such as antibiotics. NRPSs follow a modular synthetic logic whereby each successive amino-acid monomer is added to the peptide chain by successive multi-domain modules. The condensation domain catalyzes the central chemical event in the synthetic cycle, peptide-bond formation, and is present in every elongation module of the NRPS. Viomycin is an antituberculosis nonribosomal peptide that is synthesized by a series of four NRPS proteins and then modified by tailoring proteins. In order to study the mechanisms of peptide-bond formation in Viomycin and in NRPSs in general, a structural study of the first condensation domain of the Viomycin synthetase protein VioA (VioA-C1) was initiated. The gene for VioA-C1 was cloned from genomic DNA of Streptomyces vinaceus, expressed as an octahistidine-tagged construct and purified by column chromatography. VioA-C1 was crystallized using the sitting-drop vapor-diffusion method. X-ray diffraction data were collected on a rotating-anode source to 2.9 A resolution. The data could be indexed in the orthorhombic space group P212121, with unit-cell parameters a = 46.165, b = 68.335, c = 146.423 A. There is likely to be one monomer in the asymmetric unit, giving a solvent content of 49.2% and a Matthews coefficient (VM) of 2.42 A(3) Da(-1). Structural determination is in progress.

Studies of viomycin, an anti-tuberculosis antibiotic: copper(ii) coordination, DNA degradation and the impact on delta ribozyme cleavage activity.[Pubmed:27143296]

Dalton Trans. 2016 May 17;45(20):8645-58.

Viomycin is a basic peptide antibiotic, which is among the most effective agents against multidrug-resistant tuberculosis. In this paper we provide the characteristics of its acid base properties, coordination preferences towards the Cu(ii) ions, as well as the reactivity of the resulting complexes against plasmid DNA and HDV ribozyme. Careful coordination studies throughout the wide pH range allow for the characterisation of all the Cu(ii)-Viomycin complex species. The assignment of proton chemical shifts was achieved by NMR experiments, while the DTF level of theory was applied to support molecular structures of the studied complexes. The experiments with the plasmid DNA reveal that at the physiological levels of hydrogen peroxide the Cu(ii)-Viomycin complex is more aggressive against DNA than uncomplexed metal ions. Moreover, the degradation of DNA by Viomycin can be carried out without the presence of transition metal ions. In the studies of antigenomic delta ribozyme catalytic activity, Viomycin and its complex are shown to modulate the ribozyme functioning. The molecular modelling approach allows the indication of two different locations of Viomycin binding sites to the ribozyme.

Molecular mechanism of viomycin inhibition of peptide elongation in bacteria.[Pubmed:26755601]

Proc Natl Acad Sci U S A. 2016 Jan 26;113(4):978-83.

Viomycin is a tuberactinomycin antibiotic essential for treating multidrug-resistant tuberculosis. It inhibits bacterial protein synthesis by blocking elongation factor G (EF-G) catalyzed translocation of messenger RNA on the ribosome. Here we have clarified the molecular aspects of Viomycin inhibition of the elongating ribosome using pre-steady-state kinetics. We found that the probability of ribosome inhibition by Viomycin depends on competition between Viomycin and EF-G for binding to the pretranslocation ribosome, and that stable Viomycin binding requires an A-site bound tRNA. Once bound, Viomycin stalls the ribosome in a pretranslocation state for a minimum of approximately 45 s. This stalling time increases linearly with Viomycin concentration. Viomycin inhibition also promotes futile cycles of GTP hydrolysis by EF-G. Finally, we have constructed a kinetic model for Viomycin inhibition of EF-G catalyzed translocation, allowing for testable predictions of tuberactinomycin action in vivo and facilitating in-depth understanding of resistance development against this important class of antibiotics.

The antibiotic viomycin traps the ribosome in an intermediate state of translocation.[Pubmed:17515906]

Nat Struct Mol Biol. 2007 Jun;14(6):493-7.

During protein synthesis, transfer RNA and messenger RNA undergo coupled translocation through the ribosome's A, P and E sites, a process catalyzed by elongation factor EF-G. Viomycin blocks translocation on bacterial ribosomes and is believed to bind at the subunit interface. Using fluorescent resonance energy transfer and chemical footprinting, we show that Viomycin traps the ribosome in an intermediate state of translocation. Changes in FRET efficiency show that Viomycin causes relative movement of the two ribosomal subunits indistinguishable from that induced by binding of EF-G with GDPNP. Chemical probing experiments indicate that Viomycin induces formation of a hybrid-state translocation intermediate. Thus, Viomycin inhibits translation through a unique mechanism, locking ribosomes in the hybrid state; the EF-G-induced 'ratcheted' state observed by cryo-EM is identical to the hybrid state; and, since translation is Viomycin sensitive, the hybrid state may be present in vivo.

Modulation of RNA function by aminoglycoside antibiotics.[Pubmed:10619838]

EMBO J. 2000 Jan 4;19(1):1-9.

One of the most important families of antibiotics are the aminoglycosides, including drugs such as neomycin B, paromomycin, gentamicin and streptomycin. With the discovery of the catalytic potential of RNA, these antibiotics became very popular due to their RNA-binding capacity. They serve for the analysis of RNA function as well as for the study of RNA as a potential therapeutic target. Improvements in RNA structure determination recently provided first insights into the decoding site of the ribosome at high resolution and how aminoglycosides might induce misreading of the genetic code. In addition to inhibiting prokaryotic translation, aminoglycosides inhibit several catalytic RNAs such as self-splicing group I introns, RNase P and small ribozymes in vitro. Furthermore, these antibiotics interfere with human immunodeficiency virus (HIV) replication by disrupting essential RNA-protein contacts. Most exciting is the potential of many RNA-binding antibiotics to stimulate RNA activities, conceiving small-molecule partners for the hypothesis of an ancient RNA world. SELEX (systematic evolution of ligands by exponential enrichment) has been used in this evolutionary game leading to small synthetic RNAs, whose NMR structures gave valuable information on how aminoglycosides interact with RNA, which could possibly be used in applied science.

The allosteric three-site model for the ribosomal elongation cycle. New insights into the inhibition mechanisms of aminoglycosides, thiostrepton, and viomycin.[Pubmed:2843509]

J Biol Chem. 1988 Sep 15;263(26):13103-11.

According to the allosteric three-site model for the ribosomal elongation cycle (Rheinberger, H.J. and Nierhaus, K.H. (1986) J. Biol. Chem. 261, 9133-9139), two types of A site (aminoacyl-tRNA site) occupation exist. First is the A site occupation after initiation (i-type), with only one site, the P site (peptidyl-tRNA site), being prefilled with a tRNA (initiator tRNA). Second is the A site occupation after an elongation cycle (e-type), with two prefilled sites, namely the P and E sites containing peptidyl-tRNA and deacylated tRNA, respectively. The individual reactions of the elongation cycle were tested, including both types of A site occupation in the presence of various antibiotics. A test system was used allowing the functional studies to be made with quantitative tRNA binding at 6 mM Mg2+. The following results were obtained: 1) thiostrepton (5 x 10(-6) M) induced a complete block of both EF-(elongation factor) G dependent and EF-G independent translocation, in agreement with older observations. The A-site occupation of the e-type was severely inhibited in contrast to that of the i-type. Thus, thiostrepton blocks the allosteric transitions in both directions, i.e. the transition from pre- to post-translocational state (translocation) and that from the post- to the pre-translocational state (A site occupation of the e-type). In addition the ribosomal binding of EF-G.[3H] GMPPNP was inhibited by about 60%. 2) Similarly, Viomycin (5 x 10(-5) M) appears to be an inhibitor of both allosteric transitions, since it strongly inhibited the e-type (but not the i-type) A site occupation in addition to translocation. 3) The aminoglycosides streptomycin, hygromycin B, neomycin, kanamycin, and gentamicin prevented A site occupation of the e-type (residual activity below 15%). Neomycin and hygromycin, in addition, blocked the translocation reaction. Only marginal effects were observed with A site occupation of the i-type. It appears that the inhibition of the A site binding of the e-type (allosteric transition from the post- to the pretranslocational state) is the predominant effect of the misreading-inducing aminoglycosides.