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Frontiers in Microbiology 2023Aminoglycosides, as important clinical antimicrobials, are used as second-line drugs for treating multidrug-resistant tuberculosis or combined with β-lactam drugs for...
BACKGROUND
Aminoglycosides, as important clinical antimicrobials, are used as second-line drugs for treating multidrug-resistant tuberculosis or combined with β-lactam drugs for treating severe infections such as sepsis. Aminoglycoside-modifying enzyme (AME) is the most important mechanism of aminoglycoside resistance and deserves more attention.
METHODS
The bacterium DW18 was isolated from the sewage of an animal farm using the conventional method. The agar dilution method was used to determine the minimum inhibitory concentrations (MICs) of antimicrobials. A novel resistance gene was cloned, and the enzyme was expressed. The kinetic parameters were measured by a SpectraMax M5 multifunctional microplate reader. Bioinformatic analysis was performed to reveal the genetic context of the gene and its phylogenetic relationship with other AMEs.
RESULTS
A novel aminoglycoside 3'--phosphotransferase gene designated was identified in DW18 and shared the highest amino acid identity of 77.49% with the functionally characterized aminoglycoside 3'--phosphotransferase APH(3')-Ia. The recombinant plasmid carrying the novel resistance gene (pMD19-/ DH5α) showed 1,024-, 512-, 128- and 16-fold increased MIC levels for kanamycin, ribostamycin, paromomycin and neomycin, respectively, compared with the reference strain DH5α. APH(3')-Id showed the highest catalytic efficiency for ribostamycin [ of (4.96 ± 1.63) × 10 M/s], followed by paromomycin [ of (2.18 ± 0.21) × 10 M/s], neomycin [ of (1.73 ± 0.20) × 10 M/s], and kanamycin [ of (1.10 ± 0.18) × 10 M/s]. Three conserved functional domains of the aminoglycoside phosphotransferase family and ten amino acid residues responsible for the phosphorylation of kanamycin were found in the amino acid sequence of APH(3')-Id. No mobile genetic element (MGE) was discovered surrounding the gene.
CONCLUSION
In this work, a novel aminoglycoside 3'--phosphotransferase gene designated encoded in the chromosome of the environmental isolate DW18 was identified and characterized. These findings will help clinicians select effective antimicrobials to treat infections caused by pathogens with this kind of resistance gene.
PubMed: 37700861
DOI: 10.3389/fmicb.2023.1224464 -
Molecules and Cells Aug 2005A cluster of genes for ribostamycin (Rbm) biosynthesis was isolated from Streptomyces ribosidificus ATCC 21294. Sequencing of 31.892 kb of the genomic DNA of S.... (Comparative Study)
Comparative Study
A cluster of genes for ribostamycin (Rbm) biosynthesis was isolated from Streptomyces ribosidificus ATCC 21294. Sequencing of 31.892 kb of the genomic DNA of S. ribosidificus revealed 26 open reading frames (ORFs) encoding putative Rbm biosynthetic genes as well as resistance and other genes. One of ten putative Rbm biosynthetic genes, rbmA, was expressed in S. lividans TK24, and shown to encode 2-deoxy-scyllo-inosose (DOI) synthase. Acetylation of various aminoglycoside-aminocyclitol (AmAcs) by RbmI confirmed it to be an aminoglycoside 3-N-acetyltransferase. Comparison of the genetic control of ribostamycin and butirosin biosynthesis pointed to a common biosynthetic route for these compounds, despite the considerable differences between them in genetic organization.
Topics: Acetylation; Aminoglycosides; Butirosin Sulfate; Chromatography, High Pressure Liquid; Drug Resistance, Bacterial; Models, Biological; Multigene Family; Ribostamycin; Streptomyces
PubMed: 16258246
DOI: No ID Found -
The Journal of Antibiotics Dec 1983Actinomycetes were characterized in terms of resistance to 11 different aminoglycoside antibiotics (AGs). Strains freshly isolated in AG containing media showed wide... (Comparative Study)
Comparative Study
Actinomycetes were characterized in terms of resistance to 11 different aminoglycoside antibiotics (AGs). Strains freshly isolated in AG containing media showed wide varieties of multiple AG resistance, while the majority of ISP (International Streptomyces Project) cultures and the actinomycete strains isolated in an AG free medium were susceptible to all or most of the AGs tested. Marked characteristics were noted in multiple AG resistance of gray and yellow colored actinomycetes and AG-producing strains. In gray colored isolates, multiple resistance to kanamycin A, dibekacin, ribostamycin, butirosin A, istamycin A and neamine was often observed. Yellow colored isolates having multiple AG resistance were mostly resistant to neamine, ribostamycin and streptomycin and, to a lesser extent, istamycin A, dibekacin and butirosin A. Most of the AG producers tested showed unique multiple AG resistance patterns.
Topics: Actinomycetales; Aminoglycosides; Anti-Bacterial Agents; Drug Resistance, Microbial; Species Specificity; Streptomyces; Structure-Activity Relationship
PubMed: 6662815
DOI: 10.7164/antibiotics.36.1748 -
Zhongguo Yao Li Xue Bao = Acta... May 1991The experiment consists of two groups, namely the ribostamycin (Rib) group and the Rib and carbenicillin (Car) combination group. There were 7 healthy volunteers in each...
The experiment consists of two groups, namely the ribostamycin (Rib) group and the Rib and carbenicillin (Car) combination group. There were 7 healthy volunteers in each group. The Rib concentration in blood and urine was detected with the improved microbiology method. The organism tested was Bacillus pumilus. The results demonstrated that the drug serum level was in line with the one-compartment open model. The parameters such as K, Ka, T1/2, T1/2a, Tp, Cmax, and AUC in the Rib group and the combination group were 0.43 +/- 0.03 and 0.41 +/- 0.04 h-1, 3.09 +/- 1.37 and 2.82 +/- 1.48 h-1, 1.61 +/- 0.12 and 1.72 +/- 0.17 h, 0.26 +/- 0.12 and 0.31 +/- 0.15 h, 0.91 +/- 0.20 and 1.01 +/- 0.31 h, 37 +/- 4 and 34 +/- 6 micrograms.ml-1, 123 +/- 17 and 120 +/- 16 micrograms.h.ml-1, respectively. The 24-h urinary recovery rates of the 2 groups were 84 +/- 3% and 84 +/- 5%, respectively. There was no significant difference (P greater than 0.05) between the parameters of the 2 groups. The results indicate that Car has no apparent effect on the pharmacokinetics of Rib in the healthy volunteers.
Topics: Adult; Carbenicillin; Drug Therapy, Combination; Female; Humans; Male; Ribostamycin
PubMed: 1781284
DOI: No ID Found -
Biochimie May 2010Calorimetric and fluorescence techniques were used to characterize the binding of aminoglycosides-neomycin, paromomycin, and ribostamycin, with...
Calorimetric and fluorescence techniques were used to characterize the binding of aminoglycosides-neomycin, paromomycin, and ribostamycin, with 5'-dA(12)-x-dT(12)-x-dT(12)-3' intramolecular DNA triplex (x = hexaethylene glycol) and poly(dA).2poly(dT) triplex. Our results demonstrate the following features: (1) UV thermal analysis reveals that the T(m) for triplex decreases with increasing pH value in the presence of neomycin, while the T(m) for the duplex remains unchanged. (2) The binding affinity of neomycin decreases with increased pH, although there is an increase in observed binding enthalpy. (3) ITC studies conducted in two buffers (sodium cacodylate and MOPS) yield the number of protonated drug amino groups (Deltan) as 0.29 and 0.40 for neomycin and paromomycin interaction with 5'-dA(12)-x-dT(12)-x-dT(12)-3', respectively. (4) The specific heat capacity change (DeltaC(p)) determined by ITC studies is negative, with more negative values at lower salt concentrations. From 100 mM to 250 mM KCl, the DeltaC(p) ranges from -402 to -60 cal/(mol K) for neomycin. At pH 5.5, a more positive DeltaC(p) is observed, with a value of -98 cal/(mol K) at 100 mM KCl. DeltaC(p) is not significantly affected by ionic strength. (5) Salt dependence studies reveal that there are at least three amino groups of neomycin participating in the electrostatic interactions with the triplex. (6) FID studies using thiazole orange were used to derive the AC(50) (aminoglycoside concentration needed to displace 50% of the dye from the triplex) values. Neomycin shows a seven fold higher affinity than paromomycin and eleven fold higher affinity than ribostamycin at pH 6.8. (7) Modeling studies, consistent with UV and ITC results, show the importance of an additional positive charge in triplex recognition by neomycin. The modeling and thermodynamic studies indicate that neomycin binding to the DNA triplex depends upon significant contributions from charge as well as shape complementarity of the drug to the DNA triplex Watson-Hoogsteen groove.
Topics: Aminoglycosides; Calorimetry; Circular Dichroism; DNA; Hydrogen-Ion Concentration; Models, Molecular; Osmolar Concentration; Potassium Chloride; Protein Denaturation; Spectrometry, Fluorescence; Spectrophotometry, Ultraviolet
PubMed: 20167243
DOI: 10.1016/j.biochi.2010.02.004 -
The Journal of Antibiotics Aug 1982Streptomyces tenebrarius ISP 5477, which produces nebramycins, was highly resistant to the following aminoglycoside antibiotics: neamine, ribostamycin, butirosin A,...
Streptomyces tenebrarius ISP 5477, which produces nebramycins, was highly resistant to the following aminoglycoside antibiotics: neamine, ribostamycin, butirosin A, neomycin B, paromomycin, kanamycin A, dibekacin, gentamicin C complex, lividomycin A, istamycin B and streptomycin. Polyphenylalanine synthesis on the ribosomes of this strain was highly resistant to neamine, ribostamycin, butirosin A, kanamycins A, B and C, dibekacin, gentamicin C complex and istamycin B, moderately resistant to lividomycin A and streptomycin, but sensitive to neomycin B and paromomycin. Moreover, cell free extract of the strain contained phosphotransferase and N-acetyltransferase. The former enzyme was confirmed to be an aminoglycoside 6-phosphotransferase which inactivated streptomycin; the latter inactivated kanamycins B and C, dibekacin, neamine, neomycin B, paromomycin, lividomycin A, butirosin A and ribostamycin, but did not inactivate kanamycin A, gentamicin C complex and sagamicin, suggesting an aminoglycoside 2'-acetyltransferase. These results indicated that the high resistance of S. tenebrarius ISP 5477 to a wide range of aminoglycoside antibiotics is due to ribosomal resistance and to the inactivating enzymes, aminoglycoside N-acetyltransferase(s) and aminoglycoside 6-phosphotransferase.
Topics: Acetyltransferases; Aminoglycosides; Anti-Bacterial Agents; Drug Resistance, Microbial; Nebramycin; Phosphorylation; Ribosomes; Streptomyces
PubMed: 7142002
DOI: 10.7164/antibiotics.35.1020 -
ACS Infectious Diseases Oct 2019A series of derivatives of the 4,5-disubstituted class of 2-deoxystreptamine aminoglycoside antibiotics neomycin, paromomycin, and ribostamycin was prepared and assayed...
Modification at the 2'-Position of the 4,5-Series of 2-Deoxystreptamine Aminoglycoside Antibiotics To Resist Aminoglycoside Modifying Enzymes and Increase Ribosomal Target Selectivity.
A series of derivatives of the 4,5-disubstituted class of 2-deoxystreptamine aminoglycoside antibiotics neomycin, paromomycin, and ribostamycin was prepared and assayed for (i) their ability to inhibit protein synthesis by bacterial ribosomes and by engineered bacterial ribosomes carrying eukaryotic decoding A sites, (ii) antibacterial activity against wild type Gram negative and positive pathogens, and (iii) overcoming resistance due to the presence of aminoacyl transferases acting at the 2'-position. The presence of five suitably positioned residual basic amino groups was found to be necessary for activity to be retained upon removal or alkylation of the 2'-position amine. As alkylation of the 2'-amino group overcomes the action of resistance determinants acting at that position and in addition results in increased selectivity for the prokaryotic over eukaryotic ribosomes, it constitutes an attractive modification for introduction into next generation aminoglycosides. In the neomycin series, the installation of small (formamide) or basic (glycinamide) amido groups on the 2'-amino group is tolerated.
Topics: Aminoglycosides; Anti-Bacterial Agents; Bacteria; Binding Sites; Drug Resistance, Multiple, Bacterial; Hexosamines; Humans; Microbial Sensitivity Tests; Neomycin; Paromomycin; Protein Biosynthesis; Ribosomes; Structure-Activity Relationship
PubMed: 31436080
DOI: 10.1021/acsinfecdis.9b00128 -
PloS One 2013Muraymycin, a potent translocase I (MraY) inhibitor, is produced by Streptomyces sp. NRRL30471. The muraymycin gene cluster (mur) was recently cloned, and bioinformatic...
BACKGROUND
Muraymycin, a potent translocase I (MraY) inhibitor, is produced by Streptomyces sp. NRRL30471. The muraymycin gene cluster (mur) was recently cloned, and bioinformatic analysis of mur34 revealed its encoding product exhibits high homology to a large family of proteins, including KanI and RacI in individual biosynthetic pathway of kanamycin and ribostamycin. However, the precise role of these proteins remains unknown.
PRINCIPAL FINDINGS
Here we report the identification of Mur34 as the novel negative regulator involved in muraymycin biosynthesis. Independent disruption of mur34 on chromosome and cosmid directly resulted in significant improvement of muraymycin production by at least 10 folds, thereof confirming the negative function of Mur34 during muraymycin biosynthesis and realizing the engineered production of muraymycin in heterologous host. Gene expression analysis indicated that the transcription level of the mur genes in mur34 mutant (DM-5) was dramatically enhanced by ca. 30 folds. Electrophoretic mobility shift assay (EMSA) showed that Mur34 specifically bound to the promoter region of mur33. Further experiments showed that a 28-bp region downstream of the transcription start point (TSP) was protected by His6Mur34, and the -10 region is essential for the activity of mur33 promoter.
CONCLUSIONS
Mur34 plays an unambiguously negative role in muraymycin biosynthesis via binding to the upstream of mur33. More importantly, Mur34 represents a novel family of regulators acting in negative manner to regulate the secondary metabolites biosynthesis in bacteria.
Topics: Anti-Bacterial Agents; Bacterial Proteins; Base Sequence; Catechol 2,3-Dioxygenase; Computational Biology; Molecular Sequence Data; Multigene Family; Mutation; Nucleosides; Promoter Regions, Genetic; Streptomyces
PubMed: 24143177
DOI: 10.1371/journal.pone.0076068 -
Nucleic Acids Research 2006The kissing-loop complex that initiates dimerization of genomic RNA is crucial for Human Immunodeficiency Virus Type 1 (HIV-1) replication. We showed that owing to its...
The kissing-loop complex that initiates dimerization of genomic RNA is crucial for Human Immunodeficiency Virus Type 1 (HIV-1) replication. We showed that owing to its strong similitude with the bacterial ribosomal A site it can be targeted by aminoglycosides. Here, we present its crystal structure in complex with neamine, ribostamycin, neomycin and lividomycin. These structures explain the specificity for 4,5-disubstituted 2-deoxystreptamine (DOS) derivatives and for subtype A and subtype F kissing-loop complexes, and provide a strong basis for rational drug design. As a consequence of the different topologies of the kissing-loop complex and the A site, these aminoglycosides establish more contacts with HIV-1 RNA than with 16S RNA. Together with biochemical experiments, they showed that while rings I, II and III confer binding specificity, rings IV and V are important for affinity. Binding of neomycin, paromomycin and lividomycin strongly stabilized the kissing-loop complex by bridging the two HIV-1 RNA molecules. Furthermore, in situ footprinting showed that the dimerization initiation site (DIS) of HIV-1 genomic RNA could be targeted by these aminoglycosides in infected cells and virions, demonstrating its accessibility.
Topics: Aminoglycosides; Anti-HIV Agents; Binding Sites; Cell Line; Crystallography, X-Ray; Dimerization; Drug Delivery Systems; HIV-1; Humans; Models, Molecular; RNA, Viral; Virion
PubMed: 16679451
DOI: 10.1093/nar/gkl317 -
Journal of Applied Microbiology 2005To investigate the in vitro antifungal and antioomycete activities of some aminoglycosides against true fungi and Phytophthora and Pythium species and to evaluate the...
AIMS
To investigate the in vitro antifungal and antioomycete activities of some aminoglycosides against true fungi and Phytophthora and Pythium species and to evaluate the potential of the antibiotics against Phytophthora late blight on plants.
METHODS AND RESULTS
Antifungal and antioomycete activities of aminoglycoside antibiotics (neomycin, paromomycin, ribostamycin and streptomycin) and a paromomycin-producing strain (Streptomyces sp. AMG-P1) against Phytophthora and Pythium species and 10 common fungi were measured in potato dextrose broth (PDB) and on seedlings in pots. Paromomycin was the most active against Phytophthora and Pythium species with a minimal inhibitory concentration of 1-10 microg ml(-1) in PDB, but displayed low to moderate activities towards other common fungi at the same concentration. Paromomycin also showed potent in vivo activity against red pepper and tomato late blight diseases with 80 and 99% control value, respectively, at 100 microg ml(-1). In addition, culture broth of Streptomyces sp. AMG-P1 as a paromomycin producer exhibited high in vivo activity against late blight at 500 microg freeze-dried weight per millilitre.
CONCLUSIONS
Among tested aminoglycoside antibiotics, paromomycin was the most active against oomycetes both in vitro and in vivo.
SIGNIFICANCE AND IMPACT OF THE STUDY
Data from this study show that aminoglycoside antibiotics have in vitro and in vivo activities against oomycetes, suggesting that Streptomyces sp. AMG-P1 may be used as a biocontrol agent against oomycete diseases.
Topics: Antifungal Agents; Capsicum; Culture Media; Fungi; Solanum lycopersicum; Neomycin; Paromomycin; Phytophthora; Plant Diseases; Pythium; Ribostamycin; Streptomycin
PubMed: 16162234
DOI: 10.1111/j.1365-2672.2005.02684.x