Aminoglycosides Modified by Resistance Enzymes Display Diminished Binding to the Bacterial Ribosomal Aminoacyl-tRNA Site

Llano-Sotelo, Beatriz; Azucena, Eduardo; Kotra, Lakshmi P.; Mobashery, Shahriar; Chow, Christine S.

doi:10.1016/s1074-5521(02)00125-4

Cited by 155 publications

(117 citation statements)

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“…Many secondary metabolites produced by NRPSs, PKSs, and hybrid NRPS-PKS enzymes are medically important antimicrobials, immunosuppressants, or anticancer molecules (19,20). Furthermore, the cluster distal to btaR2 also harbors genes that may play a role in the immunity to and transport of antibiotics (36,40,65). We hypothesize that B. thailandensis uses BtaR2-BtaI2 quorum sensing to control antibiotic production.…”

Section: Resultsmentioning

confidence: 99%

Quorum-Sensing Control of Antibiotic Synthesis in Burkholderia thailandensis

et al. 2009

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The genome of Burkholderia thailandensis codes for several LuxR-LuxI quorum-sensing systems. We used B. thailandensis quorum-sensing deletion mutants and recombinant Escherichia coli to determine the nature of the signals produced by one of the systems, BtaR2-BtaI2, and to show that this system controls genes required for the synthesis of an antibiotic. BtaI2 is an acyl-homoserine lactone (acyl-HSL) synthase that produces two hydroxylated acyl-HSLs, N-3-hydroxy-decanoyl-HSL (3OHC 10 -HSL) and N-3-hydroxyoctanoyl-HSL (3OHC 8 -HSL). The btaI2 gene is positively regulated by BtaR2 in response to either 3OHC 10 -HSL or 3OHC 8 -HSL. The btaR2-btaI2 genes are located within clusters of genes with annotations that suggest they are involved in the synthesis of polyketide or peptide antibiotics. Stationary-phase cultures of wild-type B. thailandensis, but not a btaR2 mutant or a strain deficient in acyl-HSL synthesis, produced an antibiotic effective against gram-positive bacteria. Two of the putative antibiotic synthesis gene clusters require BtaR2 and either 3OHC 10 -HSL or 3OHC 8 -HSL for activation. This represents another example where antibiotic synthesis is controlled by quorum sensing, and it has implications for the evolutionary divergence of B. thailandensis and its close relatives Burkholderia pseudomallei and Burkholderia mallei. Burkholderia thailandensis, Burkholderia pseudomallei, andBurkholderia mallei are closely related betaproteobacteria. These three species show extensive genome sequence identity (37, 72). B. mallei is an obligate mammalian pathogen that infects solipeds (horses, mules, and donkeys) and can also cause human infections (66, 69). B. pseudomallei is an emerging human pathogen found in soil and stagnant water, and it infects rice farmers in Southeast Asia and people of Northern Australia (8,70). Once thought to be a nonpathogenic saprophyte endemic to the water and soil environments of central and northeastern Thailand (4), B. thailandensis has recently been found in the central United States and Australia (25,34). Sufficiently high doses of B. thailandensis can cause mammalian infections (4,11,56).We have been interested in acyl-homoserine lactone (acyl-HSL) quorum sensing among the B. mallei-B. pseudomallei-B. thailandensis group because quorum sensing has been implicated in the virulence of B. mallei and B. pseudomallei and because each member of this group has multiple quorumsensing systems (60-62). Quorum sensing involves production of and response to a self-produced extracellular signal that allows individuals to monitor the group population density (3,21,67). Proteobacteria often use acyl-HSL signals for quorum sensing. The signals are produced by members of the LuxI family of acyl-HSL synthases. The proteins that respond to acyl-HSLs are members of the LuxR family of transcription factors (22). Dozens of species possess acyl-HSL quorum-sensing systems. The genes coding for LuxR and LuxI homologs are often adjacent on a chromosome and are considered cognate pairs. The nature of the...

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Section: Resultsmentioning

confidence: 99%

Quorum-Sensing Control of Antibiotic Synthesis in Burkholderia thailandensis

et al. 2009

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“…1). The most important mechanism of bacterial resistance to this class of antibiotics is enzymatic modification, which greatly diminishes the affinity of aminoglycosides for the bacterial 30 S ribosomal subunit, their target in vivo (2,3). Three families of aminoglycoside-modifying enzymes are known: aminoglycoside O-nucleotidyltransferases, aminoglycoside N-acetyltransferases, and aminoglycoside O-phosphotransferases.…”

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confidence: 99%

Revisiting the Nucleotide and Aminoglycoside Substrate Specificity of the Bifunctional Aminoglycoside Acetyltransferase(6′)-Ie/Aminoglycoside Phosphotransferase(2″)-Ia Enzyme

Frase

Tóth

Vakulenko

2012

Journal of Biological Chemistry

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Background:The bifunctional AAC(6Ј)-Ie/APH(2Љ)-Ia enzyme was reported to phosphorylate all classes of aminoglycoside antibiotics using ATP. Results: GTP, and not ATP, is the cosubstrate of the enzyme. 4,5-disubstituted and atypical aminoglycosides are not substrates. Conclusion:The enzyme is a narrow spectrum GTP-dependent kinase that phosphorylates 4,6-disubstituted aminoglycosides exclusively. Significance: Knowledge of enzyme activity is essential for developing novel antibiotics and conducting effective antimicrobial therapy.

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“…1 and 2). Bacteria become protected from aminoglycosides, because the modified antibiotics can no longer bind with high affinity to their target, the A-site of the small ribosomal subunit, because of unfavorable steric and/or electrostatic constraints (3). The AMEs are a diverse set of proteins composed of three families: aminoglycoside nucleotidyltransferases, aminoglycoside acetyltransferases (AACs), and aminoglycoside phosphotransferases (APHs).…”

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confidence: 99%

“…3 Its inability to inactivate AAC(6Ј)-Ii could be because 1-(bromomethyl)phenanthrene can not efficiently bind to AAC(6Ј)-Ii or because AAC(6Ј)-Ii lacks an appropriate active site residue to serve as the nucleophile in the reaction.…”

mentioning

confidence: 99%

The Molecular Basis of the Expansive Substrate Specificity of the Antibiotic Resistance Enzyme Aminoglycoside Acetyltransferase-6′-Aminoglycoside Phosphotransferase-2“

Boehr

Jenkins

Wright

2003

Journal of Biological Chemistry

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The most frequent determinant of aminoglycoside antibiotic resistance in Gram-positive bacterial pathogens is a bifunctional enzyme, aminoglycoside acetyltransferase-6 -aminoglycoside phosphotransferase-2؆ (AAC(6 )-aminoglycoside phosphotransferase-2؆, capable of modifying a wide selection of clinically relevant antibiotics through its acetyltransferase and kinase activities. The aminoglycoside acetyltransferase domain of the enzyme, AAC(6 )-Ie, is the only member of the large AAC(6 ) subclass known to modify fortimicin A and catalyze Oacetylation. We have demonstrated through solvent isotope, pH, and site-directed mutagenesis effects that Asp-99 is responsible for the distinct abilities of AAC(6 )-Ie. Moreover, we have demonstrated that small planar molecules such as 1-(bromomethyl)phenanthrene can inactivate the enzyme through covalent modification of this residue. Thus, Asp-99 acts as an active site base in the molecular mechanism of AAC(6 )-Ie. The prominent role of this residue in aminoglycoside modification can be exploited as an anchoring site for the development of compounds capable of reversing antibiotic resistance in vivo.Clinical usage of aminoglycoside-aminocylitol antibiotics is blocked by the presence of aminoglycoside modifying enzymes (AMEs) 1 in resistant organisms (for review see Refs. 1 and 2). Bacteria become protected from aminoglycosides, because the modified antibiotics can no longer bind with high affinity to their target, the A-site of the small ribosomal subunit, because of unfavorable steric and/or electrostatic constraints (3). The AMEs are a diverse set of proteins composed of three families: aminoglycoside nucleotidyltransferases, aminoglycoside acetyltransferases (AACs), and aminoglycoside phosphotransferases (APHs).The most clinically important AME in Gram-positive bacterial pathogens such as Staphylococcus aureus is AAC(6Ј)-APH(2Љ) (4). This enzyme is unique in that it is bifunctional, comprising activities from two classes of AMEs: an N-terminal AAC (AAC(6Ј)-Ie) and a C-terminal APH (APH(2Љ)-Ia) ( Fig. 1) (5, 6). AAC(6Ј)-APH(2Љ) has an extraordinary ability to detoxify a wide selection of aminoglycosides (7, 8) not only as a result of its bifunctional nature but also because of the special characteristics of each activity. For example, APH(2Љ)-Ia has a tremendous ability to accommodate a vast range of antibiotics and binding conformations as evidenced by the remarkably broad regiospecificity of phosphoryl transfer that enables modification to occur on hydroxyl groups from four different aminoglycoside ring systems (see Fig. 1) (8). Conversely, AAC(6Ј)-Ie, has a very stringent regiospecificity of acetyl transfer, because it is restricted to acetylation of the 6Ј-position of aminoglycosides only (see Fig. 1) (8). AAC(6Ј)-Ie is the sole member of the very large AAC(6Ј) subclass known to acetylate the antibiotic fortimicin A (9), and moreover, in addition to N-acetylation activity, this enzyme has been shown to have O-acetylation capabilities (8). The unusual activities of AAC(6Ј)-...

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Aminoglycosides Modified by Resistance Enzymes Display Diminished Binding to the Bacterial Ribosomal Aminoacyl-tRNA Site

Cited by 155 publications

References 41 publications

Quorum-Sensing Control of Antibiotic Synthesis in Burkholderia thailandensis

Quorum-Sensing Control of Antibiotic Synthesis in Burkholderia thailandensis

Revisiting the Nucleotide and Aminoglycoside Substrate Specificity of the Bifunctional Aminoglycoside Acetyltransferase(6′)-Ie/Aminoglycoside Phosphotransferase(2″)-Ia Enzyme

The Molecular Basis of the Expansive Substrate Specificity of the Antibiotic Resistance Enzyme Aminoglycoside Acetyltransferase-6′-Aminoglycoside Phosphotransferase-2“

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