Influence of Linker Length and Composition on Enzymatic Activity and Ribosomal Binding of Neomycin Dimers

Watkins, Davita L.; Kumar, Sunil; Green, Keith D.; Arya, Dev P.

doi:10.1128/aac.00861-15

Cited by 19 publications

(21 citation statements)

References 30 publications

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“…It is important to note that we did not test our NEO dimers 6-15 against ANTs due to the facts that (i) most ANTs modify streptomycin or spectinomycin, 7 and (ii) the bacterial strains against which these compounds were tested in this study are not known to contain ANTs that modify NEO. 13 …”

Section: Resultsmentioning

confidence: 99%

“…We report the synthesis and screening of a series of neomycin B (NEO) dimers that are joined by L-arginine-containing linkers of varying lengths. NEO dimers with triazole, urea, and thiourea linkages have been shown to be poor substrates for certain AMEs, 13 and both NEO dimers and L-arginine-conjugated AGs have been reported to increase ligand affinity for nucleic acids as compared with their unconjugated counterparts. 14–21 Other AG homo- and hetero-dimers have also been reported to display promise in enhancing RNA binding, improving antibiotic activity, and/or resisting the action of AMEs.…”

Section: Introductionmentioning

confidence: 99%

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Arginine-linked neomycin B dimers: synthesis, rRNA binding, and resistance enzyme activity

et al. 2016

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The nucleotides comprising the ribosomal decoding center are highly conserved, as they are important for maintaining translational fidelity. The bacterial A-site has a small base variation as compared with the human analogue, allowing aminoglycoside (AG) antibiotics to selectively bind within this region of the ribosome and negatively affect microbial protein synthesis. Here, by using a fluorescence displacement screening assay, we demonstrate that neomycin B (NEO) dimers connected by L-arginine-containing linkers of varying length and composition bind with higher affinity to model A-site RNAs compared to NEO, with IC50 values ranging from ~40–70 nM, and that a certain range of linker lengths demonstrates a clear preference for the bacterial A-site RNA over the human analogue. Furthermore, AG-modifying enzymes (AMEs), such as AG O-phosphotransferases, which are responsible for conferring antibiotic resistance in many types of infectious bacteria, demonstrate markedly reduced activity against several of the L-arginine-linked NEO dimers in vitro. The antimicrobial activity of these dimers against several bacterial strains is weaker than that of the parent NEO.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Arginine-linked neomycin B dimers: synthesis, rRNA binding, and resistance enzyme activity

et al. 2016

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“…We used NEO as a control. All AME enzymes, AAC(6′)-Ie, 47 AAC(3)-IV, 47 AAC(2′)-Ic, 48 Eis, 48 APH(2″)-Ia, 25 and APH(3′)-Ia, 24 were purified and tested as previously described. All reactions were monitored at 25 °C (with the exception of AAC(6′)-Ie and APH(2″)-Ia, which were monitored at 37 °C) on a SpectraMax M5 microplate reader and performed in duplicate.…”

Section: Methodsmentioning

confidence: 99%

“…23 We have previously demonstrated that conjugation of aminoglycosides to small molecules renders them poor substrates for AMEs. 13,14,24,25 Here, we examine 12 new anthraquinone–NEO conjugates (AMA–NEO) ( 1 – 12 ) with thiourea linkers for their antibacterial properties, translation inhibition, resistance to AMEs, and binding selectivity for five different 27-nucleotide RNA hairpin constructs representing A-site homologues (Figures 1 and 2). The Escherichia coli A-site is a highly conserved region for aminoglycoside binding in the bacterial ribosome.…”

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Antimicrobial Activity, AME Resistance, and A-Site Binding Studies of Anthraquinone–Neomycin Conjugates

et al. 2017

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The antibacterial effects of aminoglycosides are based on their association with the A-site of bacterial rRNA and interference with the translational process in the bacterial cell, causing cell death. The clinical use of aminoglycosides is complicated by resistance and side effects, some of which arise from their interactions with the human mitochondrial 12S rRNA and its deafness-associated mutations, C1494U and A1555G. We report a rapid assay that allows screening of aminoglycoside compounds to these classes of rRNAs. These screening tools are important to find antibiotics that selectively bind to the bacterial A-site rather than human, mitochondrial A-sites and its mutant homologues. Herein, we report our preliminary work on the optimization of this screen using 12 anthraquinone–neomycin (AMA–NEO) conjugates against molecular constructs representing five A-site homologues, Escherichia coli, human cytosolic, mitochondrial, C1494U, and A1555G, using a fluorescent displacement screening assay. These conjugates were also tested for inhibition of protein synthesis, antibacterial activity against 14 clinically relevant bacterial strains, and the effect on enzymes that inactivate aminoglycosides. The AMA–NEO conjugates demonstrated significantly improved resistance against aminoglycoside-modifying enzymes (AMEs), as compared with NEO. Several compounds exhibited significantly greater inhibition of prokaryotic protein synthesis as compared to NEO and were extremely poor inhibitors of eukaryotic translation. There was significant variation in antibacterial activity and MIC of selected compounds between bacterial strains, with Escherichia coli, Enteroccocus faecalis, Citrobacter freundii, Shigella flexneri, Serratia marcescens, Proteus mirabilis, Enterobacter cloacae, Staphylococcus epidermidis, and Listeria monocytogenes exhibiting moderate to high sensitivity (50–100% growth inhibition) whereas Acinetobacter baumannii, Pseudomonas aeruginosa, Klebsiellla pneumoniae, and MRSA strains expressed low sensitivity, as compared to the parent aminoglycoside NEO.

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“…In most cases, the overall yields of syntheses are rather low, requiring large quantities of starting materials and many steps to achieve a single product. With the challenges in synthesizing AGs from scratch, several approaches have targeted modifying currently approved AGs including modifications at various positions such as the 1-, 6′-, 21 2″-, 22 5″-, 23 and 6″-positions, 3,24,25 dimerization, 26,27 and covalent attachment to other antibiotics, 23,28 which have all had varying degree of success. Herein, we report on the facile syntheses of AMK, kanamycin A (KAN), netilmicin (NET), sisomicin (SIS), and tobramycin (TOB) mono-and dimodified at the 1-, 6′-, and/or 4‴-amines by glycinyl, carboxybenzyl, and AHB moieties (Scheme 1).…”

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confidence: 99%

Synthesis and Biological Activity of Mono- and Di-N-acylated Aminoglycosides

Chandrika

Green

Houghton

2015

ACS Med. Chem. Lett.

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Despite issues with oto/nephrotoxicity and bacterial resistance, aminoglycosides (AGs) remain an effective and widely used class of antibacterial agents. For decades now, efforts toward the development of novel AGs with potential to overcome some of these problems have been major research focuses. 1-N-Acylation, especially γ-amino-β-hydroxybutyrate (AHB) derivatization, has proven to be one of the most successful strategies for improving the overall properties of AGs, including their ability to avoid certain resistance mechanisms. More recently, 6′-N-acylation arose as another possible strategy to improve the properties of these drugs. In this study, we report on the glycinyl, carboxybenzyl, and AHB mono-and diderivatization at the 1-, 6′-, and/or 4‴-amines of the AGs amikacin, kanamycin A, netilmicin, sisomicin, and tobramycin. We also present the antibacterial activities and the reduced reactivity of AG-modifying enzymes (AMEs) toward these new AG derivatives, and identify the AMEs present in the bacterial strains tested.

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Influence of Linker Length and Composition on Enzymatic Activity and Ribosomal Binding of Neomycin Dimers

Cited by 19 publications

References 30 publications

Arginine-linked neomycin B dimers: synthesis, rRNA binding, and resistance enzyme activity

Arginine-linked neomycin B dimers: synthesis, rRNA binding, and resistance enzyme activity

Antimicrobial Activity, AME Resistance, and A-Site Binding Studies of Anthraquinone–Neomycin Conjugates

Synthesis and Biological Activity of Mono- and Di-N-acylated Aminoglycosides

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