Mechanistic characterization of the bifunctional aminoglycoside-modifying enzyme AAC(3)-Ib/AAC(6')-Ib' from Pseudomonas aeruginosa

A recently discovered bifunctional antibiotic-resistance enzyme named AAC(3)-Ib/AAC(6')-Ib', from Pseudomonas aeruginosa, catalyzes acetylation of aminoglycoside antibiotics. Since both domains are acetyltransferases, each was cloned and purified for mechanistic studies. The AAC(3)-Ib domain appears to be highly specific to fortimicin A and gentamicin as substrates, while the AAC(6')-Ib' domain exhibits a broad substrate spectrum. Initial velocity patterns indicate that both domains follow a sequential kinetic mechanism. The use of dead-end and product inhibition and solvent-isotope effect reveals that both domains catalyze their reactions by a steady-state ordered Bi-Bi kinetic mechanism, in which acetyl-CoA is the first substrate that binds to the active site, followed by binding of the aminoglycoside antibiotic. Subsequent to the transfer of the acetyl group, acetylated aminoglycoside is released prior to coenzyme A. The merger of two genes to create a bifunctional enzyme with expanded substrate profile would appear to be a recent trend in evolution of resistance to aminoglycoside antibiotics, of which four examples have been documented in the past few years.     

Kinetic and mutagenic characterization of the chromosomally encoded Salmonella enterica AAC(6')-Iy aminoglycoside N-acetyltransferase

The chromosomally encoded aminoglycoside N-acetyltransferase, AAC(6')-Iy, from Salmonella enterica confers resistance toward a number of aminoglycoside antibiotics. The structural gene was cloned and expressed and the purified enzyme existed in solution as a dimer of ca. 17 000 Da monomers. Acetyl-CoA was the preferred acyl donor, and most therapeutically important aminoglycosides were substrates for acetylation. Exceptions are those aminoglycosides that possess a 6'-hydroxyl substituent (e.g., lividomycin). Thus, the enzyme exhibited regioselective and exclusive acetyltransferase activity to 6'-amine-containing aminoglycosides. The enzyme exhibited Michaelis-Menten kinetics for some aminoglycoside substrates but "substrate activation" with others. Kinetic studies supported a random kinetic mechanism for the enzyme. The enzyme was inactivated by iodoacetamide in a biphasic manner, with half of the activity being lost rapidly and the other half more slowly. Tobramycin, but not acetyl-CoA, protected against inactivation. Each of the three cysteine residues (C70, C109, C145) in the wild-type enzyme were carboxamidomethylated by iodoacetamide. Cysteine 109 in AAC(6')-Iy is conserved in 12 AAC(6') enzyme sequences of the major class I subfamily. Surprisingly, mutation of this residue to alanine neither abolished activity nor altered the biphasic inactivation by iodoacetamide. The maximum velocity and V/K values for a number of aminoglycosides were elevated in this single mutant, and the kinetic behavior of substrates exhibiting linear vs nonlinear kinetics was reversed. Cysteine 70 in AAC(6')-Iy is either a cysteine or a threonine residue in all 12 AAC(6') enzymes of the major class I subfamily. The double mutant, C109A/C70A, was not inactivated by iodoacetamide. The double mutant exhibited large increases in the K(m) values for both acetyl-CoA and aminoglycoside substrates, and all aminoglycoside substrates exhibited Michaelis-Menten kinetics. Solvent kinetic isotope effects on V/K were normal for the WT enzyme and inverse for the double mutant. We discuss a chemical mechanism and the likely rate-limiting steps for both the wild-type and mutant forms of the enzyme.     

Overexpression and mechanistic analysis of chromosomally encoded aminoglycoside 2'-N-acetyltransferase (AAC(2')-Ic) from Mycobacterium tuberculosis

The chromosomally encoded aminoglycoside N-acetyltransferase, AAC(2')-Ic, of Mycobacterium tuberculosis has a yet unidentified physiological function. The aac(2')-Ic gene was cloned and expressed in Escherichia coli, and AAC(2')-Ic was purified. Recombinant AAC(2')-Ic was a soluble protein of 20,000 Da and acetylated all aminoglycosides substrates tested in vitro, including therapeutically important antibiotics. Acetyl-CoA was the preferred acyl donor. The enzyme, in addition to acetylating aminoglycosides containing 2'-amino substituents, also acetylated kanamycin A and amikacin that contain a 2'-hydroxyl substituent, although with lower activity, indicating the capacity of the enzyme to perform both N-acetyl and O-acetyl transfer. The enzyme exhibited "substrate activation" with many aminoglycoside substrates while exhibiting Michaelis-Menten kinetics with others. Kinetic studies supported a random kinetic mechanism for AAC(2')-Ic. Comparison of the kinetic parameters of different aminoglycosides suggested that their hexopyranosyl residues and, to a lesser extent, the central aminocyclitol residue carry the major determinants of substrate affinity.     

Thermodynamics of aminoglycoside and acyl-coenzyme A binding to the Salmonella enterica AAC(6')-Iy aminoglycoside N-acetyltransferase

Kinetic and mechanistic studies on the chromosomally encoded aminoglycoside 6'-N-acetyltransferase, AAC(6')-Iy, of Salmonella enterica that confers resistance toward aminoglycosides have been previously reported [Magnet et al. (2001) Biochemistry 40, 3700-3709]. In the present study, equilibrium binding and the thermodynamic parameters of binding of aminoglycosides and acyl-coenzyme A derivatives to AAC(6')-Iy and of two mutants, C109A and the C109A/C70A double mutant, have been studied using fluorescence spectroscopy and isothermal titration calorimetry (ITC). Association constants for different aminoglycosides varied greatly (4 x 10(4)-150 x 10(4)) while the association constants of several acyl-coenzyme A derivatives were similar (3.2 x 10(4)-4.5 x 10(4)). The association constants and van't Hoff enthalpy changes derived from intrinsic protein fluorescence changes were in agreement with independently measured values from isothermal titration calorimetry studies. Binding of both aminoglycosides and acyl-coenzyme A derivatives is strongly enthalpically driven and revealed opposing negative entropy changes, resulting in enthalpy-entropy compensation. The acetyltransferase exhibited a temperature-dependent binding of tobramycin with a negative heat capacity value of 410 cal mol(-1) K(-1). Isothermal titration studies of acetyl-coenzyme A and tobramycin binding to mutant forms of the enzyme indicated that completely conserved C109 does not play any direct role in the binding of either of the substrates, while C70 is directly involved in aminoglycoside binding. These results are discussed and compared with previous steady-state kinetic studies of the enzyme.     

Identification of strain harboring both aac(6')-Ib and aac(6')-Ib-cr variant simultaneously in Escherichia coli and Klebsiella pneumoniae

The aac(6')-Ib gene is the most prevalent gene that encodes aminoglycoside-modifying enzymes and confers resistance to tobramycin, kanamycin, and amikacin. The aac(6')-Ib-cr variant gene can induce resistance against aminoglycoside and fluoroquinolone simultaneously. Two main methods, sequence analysis and the restriction enzyme method, can detect the aac(6')-Ib-cr variant in clinical strains. We collected the 85 strains that were believed to be aac(6')-Ib positive from clinical isolates. Among them, 38 strains were the wild-type; the remaining 47 strains were the aac(6')-Ib-cr variant. Of these 47 strains, 19 simultaneously harbored aac(6')-Ib and aac(6')-Ib-cr. Our study aims to report the characteristics of the 19 strains that simultaneously harbored both genes. This study is the first investigation published in Korea of strains that included both aac(6')-Ib and aac(6')-Ib-cr variant.     

Domain-domain interactions in the aminoglycoside antibiotic resistance enzyme AAC(6')-APH(2'')

The most common determinant of aminoglycoside antibiotic resistance in Gram positive bacterial pathogens, such as Staphylococcus aureus, is a modifying enzyme, AAC(6')-APH(2' '), capable of acetylating and phosphorylating a wide range of antibiotics. This enzyme is unique in that it is composed of two separable modification domains, and although a number of studies have been conducted on the acetyltransferase and phosphotransferase activities in isolation, little is known about the role and impact of domain interactions on antibiotic resistance. Kinetic analysis and in vivo assessment of a number of N- and C-terminal truncated proteins have demonstrated that the two domains operate independently and do not accentuate one another's resistance activity. However, the two domains are structurally integrated, and mutational analysis has demonstrated that a predicted connecting alpha-helix is especially critical for maintaining proper structure and function of both activities. AAC(6')-APH(2' ') detoxifies a staggering array of aminoglycosides, where one or both activities make important contributions depending on the antibiotic. Thus, to overcome antibiotic resistance associated with AAC(6')-APH(2' '), aminoglycosides resistant to modification and/or inhibitors against both activities must be employed. Domain-domain interactions in AAC(6')-APH(2' ') offer a unique target for inhibitor strategies, as we show that their disruption simultaneously inhibits both activities >90%.     

Characterization of the bifunctional aminoglycoside-modifying enzyme ANT(3')-Ii/AAC(6')-IId from Serratia marcescens

A newly discovered bifunctional antibiotic resistance enzyme from Serratia marcescens catalyzes adenylation and acetylation of aminoglycoside antibiotics. The structure assignment of the enzymic products indicated that acetylation takes place on the 6'-amine of kanamycin A and the adenylation on 3'- and 9-hydroxyl groups of streptomycin and spectinomycin, respectively. The adenyltransferase domain appears to be highly specific to spectinomycin and streptomycin, while the acetyltransferase domain shows a broad substrate profile. Initial velocity patterns indicate that both domains follow a sequential kinetic mechanism. The use of dead-end and product inhibition, the solvent isotope effect, and the solvent viscosity effect reveals that the adenyltransferase domain catalyzes the reaction by a Theorell-Chance kinetic mechanism, where ATP binds to the enzyme prior to the aminoglycoside and the modified antibiotic is the last product to be released. The acetyltransferase domain follows an ordered bi-bi kinetic mechanism, in which the antibiotic is the first substrate that binds to the active site and CoASH is released prior to the modified aminoglycoside. The merging of two genes to create bifunctional resistance enzymes with expanded profiles has now been documented in four instances, including the subject of study in this report, which suggests a new trend in the emergence of resistance to aminoglycoside antibiotics among pathogens.     

Characteristics of Klebsiella pneumoniae harboring QnrB32, Aac(6')-Ib-cr, GyrA and CTX-M-22 genes

Quinolone resistance in members of the Enterobacteriaceae family is mostly due to mutations in the quinolone resistance-determining regions of topoisomerase genes. CTX-M-22 is a member of the CTX-M family which can reduce extended-spectrum β-lactamase (ESBL) production and modulate antibiotic resistance, resulting in low ceftazidime minimum inhibitory concentrations (MICs). There are four different genes in Klebsiella pneumoniae (KP4707) including qnrB32 (novel qnr allele gene, HQ704413), aac(6')-Ib-cr (novel aac(6')-Ib allele gene, HQ680690), gyrA (novel gyrA allele gene, HQ680691) and CTX-M-22 gene. Five point amino acid mutations Arn(N)27 → Leu(L), Val(V)129 → Ala(A), Iie(I)142 → Met(M), Gly(G)188 → Arg(R), Val(V)212 → Iie(I) were observed in the qnr32 gene when compared to qnrB1. Of all qnrB alleles, a novel variant of the qnrB32 gene, with qnrB31, had the highest amino acid homology. Three point amino acid mutations including Trp(W)105 → Arg(R), Asp(D)182 → Tyr(Y) and Val(V)201 → Asp(D) were observed in aac(6')-Ib-cr gene, when compared to GenBank number AF479774. New variants of qnr32, aac(6')-Ib-cr, gyrA and CTX-M-22 or other genotype determinants continuously appear in different genomic sites and also outside the Enterobacteriaceae family.     

Kinetic mechanism of the GCN5-related chromosomal aminoglycoside acetyltransferase AAC(6')-Ii from Enterococcus faecium: evidence of dimer subunit cooperativity

The aminoglycoside 6'-N-acetyltransferase AAC(6')-Ii from Enterococcus faecium is an important microbial resistance determinant and a member of the GCN5-related N-acetyltransferase (GNAT) superfamily. We report here the further characterization of this enzyme in terms of the kinetic mechanism of acetyl transfer and identification of rate-contributing step(s) in catalysis, as well as investigations into the binding of both acetyl-CoA and aminoglycoside substrates to the AAC(6')-Ii dimer. Product and dead-end inhibition studies revealed that AAC(6')-Ii follows an ordered bi-bi ternary complex mechanism with acetyl-CoA binding first followed by antibiotic. Solvent viscosity studies demonstrated that aminoglycoside binding and product release govern the rate of acetyl transfer, as evidenced by changes in both the k(cat)/K(b) for aminoglycoside and k(cat), respectively, with increasing solvent viscosity. Solvent isotope effects were consistent with our viscosity studies that diffusion-controlled processes and not the chemical step were rate-limiting in drug modification. The patterns of partial and mixed inhibition observed during our mechanistic studies were followed up by investigating the possibility of subunit cooperativity in the AAC(6')-Ii dimer. Through the use of AAC-Trp(164) --> Ala, an active mutant which exists as a monomer in solution, the partial nature of the competitive inhibition observed in wild-type dead-end inhibition studies was alleviated. Isothermal titration calorimetry studies also indicated two nonequivalent antibiotic binding sites for the AAC(6')-Ii dimer but only one binding site for the Trp(164) --> Ala mutant. Taken together, these results demonstrate subunit cooperativity in the AAC(6')-Ii dimer, with possible relevance to other oligomeric members of the GNAT superfamily.     

Development of an immunochromatographic assay for rapid detection of AAC(6')-Ib-producing Pseudomonas aeruginosa

To detect aminoglycoside 6'-N-acetyltransferase-Ib [AAC(6')-Ib]-producing, Pseudomonas aeruginosa isolates which are a frequent cause of nosocomial infections in Japan, an immunochromatographic assay was developed using two kinds of monoclonal antibodies (mAbs) recognizing AAC(6')-Ib. The results of the assessment were fully consistent with those of aac(6')-Ib PCR analyses.     

Crystal structure of an aminoglycoside 6'-N-acetyltransferase: defining the GCN5-related N-acetyltransferase superfamily fold.

BACKGROUND: The predominant mechanism of antibiotic resistance employed by pathogenic bacteria against the clinically used aminoglycosides is chemical modification of the drug. The detoxification reactions are catalyzed by enzymes that promote either the phosphorylation, adenylation or acetylation of aminoglycosides. Structural studies of these aminoglycoside-modifying enzymes may assist in the development of therapeutic agents that could circumvent antibiotic resistance. In addition, such studies may shed light on the development of antibiotic resistance and the evolution of different enzyme classes. RESULTS: The crystal structure of the aminoglycoside-modifying enzyme aminoglycoside 6'-N-acetyltransferase type li (AAC(6')-li) in complex with the cofactor acetyl coenzyme A has been determined at 2.7 A resolution. The structure establishes that this acetyltransferase belongs to the GCN5-related N-acetyltransferase superfamily, which includes such enzymes as the histone acetyltransferases GCN5 and Hat1. CONCLUSIONS: Comparison of the AAC(6')-li structure with the crystal structures of two other members of this superfamily, Serratia marcescens aminoglycoside 3-N-acetyltransferase and yeast histone acetyltransferase Hat1, reveals that of the 84 residues that are structurally similar, only three are conserved and none can be implicated as catalytic residues. Despite the negligible sequence identity, functional studies show that AAC(6')-li possesses protein acetylation activity. Thus, AAC(6')-li is both a structural and functional homolog of the GCN5-related histone acetyltransferases.     

X-ray structure of the AAC(6')-Ii antibiotic resistance enzyme at 1.8 A resolution; examination of oligomeric arrangements in GNAT superfamily members

The rise of antibiotic resistance as a public health concern has led to increased interest in studying the ways in which bacteria avoid the effects of antibiotics. Enzymatic inactivation by several families of enzymes has been observed to be the predominant mechanism of resistance to aminoglycoside antibiotics such as kanamycin and gentamicin. Despite the importance of acetyltransferases in bacterial resistance to aminoglycoside antibiotics, relatively little is known about their structure and mechanism. Here we report the three-dimensional atomic structure of the aminoglycoside acetyltransferase AAC(6')-Ii in complex with coenzyme A (CoA). This structure unambiguously identifies the physiologically relevant AAC(6')-Ii dimer species, and reveals that the enzyme structure is similar in the AcCoA and CoA bound forms. AAC(6')-Ii is a member of the GCN5-related N-acetyltransferase (GNAT) superfamily of acetyltransferases, a diverse group of enzymes that possess a conserved structural motif, despite low sequence homology. AAC(6')-Ii is also a member of a subset of enzymes in the GNAT superfamily that form multimeric complexes. The dimer arrangements within the multimeric GNAT superfamily members are compared, revealing that AAC(6')-Ii forms a dimer assembly that is different from that observed in the other multimeric GNAT superfamily members. This different assembly may provide insight into the evolutionary processes governing dimer formation.     

The kinetic mechanism of AAC3-IV aminoglycoside acetyltransferase from Escherichia coli

The aminoglycoside 3-N-acetyltransferase AAC(3)-IV from Escherichia coli exhibits a very broad aminoglycoside specificity, causing resistance to a large number of aminoglycosides, including the atypical veterinary antibiotic, apramycin. We report here on the characterization of the substrate specificity and kinetic mechanism of the acetyl transfer reaction catalyzed by AAC(3)-IV. The steady-state kinetic parameters revealed a narrow specificity for the acyl-donor and broad range of activity for aminoglycosides. AAC(3)-IV has the broadest substrate specificity of all AAC(3)'s studied to date. Dead-end inhibition and ITC experiments revealed that AAC(3)-IV follows a sequential, random bi-bi kinetic mechanism. The analysis of the pH dependence of the kinetic parameters revealed acid- and base-assisted catalysis and the existence of three additional ionizable groups involved in substrate binding. The magnitude of the solvent kinetic isotope effects suggests that a chemical step is at least partially rate limiting in the overall reaction.     

Prevalence of AA(6')-APH(2")Ia gene among high-level aminoglycoside resistant Enterococci and Staphylococci

According to new reports the AAC (6')-APH (2")Ia gene is no longer the only gene encoding resistance to gentamycin in Gram-positive cocci and therefore the current method for predicting synergism aminoglycosides with bacterial cell wall active agents in this bacteria may need revision. To further our knowledge of aminoglycoside resistance mechanism in Gram-positive cocci in Gdańsk region we tested presence of AAC (6')-APH (2")Ia gene among 22 enterococcal (E. faecalis) and 41 staphylococcal (S. haemolyticus, S. aureus, S. epidermidis) gentamycin-resistant isolates. Presence of AAC (6')-APH (2")Ia gene varied from 50% (n = 6) in gentamycin-resistant S. epidermidis, 80% (n = 10) in gentamycin resistant S. haemolyticus 88% in methicillin-resistant Staphylococcus aureus (MRSA) (n = 25). In Enterococcus faecalis this gene was noticed only in 59% (n = 22) of gentamycin-resistant isolates. These results suggest that spread of resistance gene among different species is limited and AAC (6')-APH (2")Ia mediated gentamycin-resistance mechanism is more common among MRSA and Staphylococcus haemolyticus.     

Aminoglycoside 2'-N-acetyltransferase from Mycobacterium tuberculosis in complex with coenzyme A and aminoglycoside substrates

AAC(2')-Ic catalyzes the coenzyme A (CoA)-dependent acetylation of the 2' hydroxyl or amino group of a broad spectrum of aminoglycosides. The crystal structure of the AAC(2')-Ic from Mycobacterium tuberculosis has been determined in the apo enzyme form and in ternary complexes with CoA and either tobramycin, kanamycin A or ribostamycin, representing the first structures of an aminoglycoside acetyltransferase bound to a drug. The overall fold of AAC(2')-Ic places it in the GCN5-related N-acetyltransferase (GNAT) superfamily. Although the physiological function of AAC(2')-Ic is uncertain, a structural analysis of these high-affinity aminoglycoside complexes suggests that the enzyme may acetylate a key biosynthetic intermediate of mycothiol, the major reducing agent in mycobacteria, and participate in the regulation of cellular redox potential.     

Kinetic and structural analysis of bisubstrate inhibition of the Salmonella enterica aminoglycoside 6'-N-acetyltransferase

Aminoglycosides are antibacterial compounds that act by binding to the A site of the small 30S bacterial ribosomal subunit and inhibiting protein translation. Clinical resistance to aminoglycosides is generally the result of the expression of enzymes that covalently modify the antibiotic, including phosphorylation, adenylylation, and acetylation. Bisubstrate analogs for the aminoglycoside N-acetyltransferases are nanomolar inhibitors of Enterococcus faecium AAC(6')-Ii. However, in the case of the Salmonella enterica aac(6')-Iy-encoded aminoglycoside N-acetyltransferase, we demonstrate that a series of bisubstrate analogs are only micromolar inhibitors. In contrast to studies with AAC(6')-Ii, the inhibition constants toward AAC(6')-Iy are essentially independent of both the identity of the aminoglycoside component of the bisubstrate and the number of carbon atoms that are used to link the CoA and aminoglycoside components. The patterns of inhibition suggest that the CoA portion of the bisubstrate analog can bind to the enzyme-aminoglycoside substrate complex and that the aminoglycoside portion can bind to the enzyme-CoA product complex. However, at the high concentrations of bisubstrate analog used in crystallization experiments, we could crystallize and solve the three-dimensional structure of the enzyme-bisubstrate complex. The structure reveals that both the CoA and aminoglycoside portions bind in essentially the same positions as those previously observed for the enzyme-CoA-ribostamycin complex, with only a modest adjustment to accommodate the "linker". These results are compared to previous studies of the interaction of similar bisubstrate analogs with other aminoglycoside N-acetyltransferases.     

Primary structure of an aminoglycoside 6''-N-acetyltransferase AAC(6'')-4, fused in vivo with the signal peptide of the Tn3-encoded beta-lactamase.

The gene aacA4 encoding an aminoglycoside 6'-N-acetyltransferase, AAC(6')-4, was cloned from a natural multiresistance plasmid, and its nucleotide sequence was determined. The gene was 600 base pairs (bp) long, and the AAC(6')-4 had a calculated molecular size of 22.4 kilodaltons and an isoelectric point of 5.35. The sequence of the 17 N-terminal amino acids was determined from the purified enzyme. The AAC(6')-4 gene was part of a resistance gene cluster, and its expression was under the control of the regulatory sequences of the beta-lactamase encoded by Tn3. The five N-terminal amino acids were identical to those of the signal peptide of the Tn3-encoded beta-lactamase, and the entire 5' region of aacA4, as far as it was sequenced (354 bp, including the promoter and the ribosome-binding site sequences), was identical to that of the beta-lactamase gene. This led us to presume an in vivo fusion between the beta-lactamase and the acetyltransferase genes. The latter was followed, in a polycistronic arrangement, by an aminoglycoside 3",9-adenylyltransferase gene, aadA, with an intergenic region of 68 bp. At a distance of ca. 1.3 kilobases in the 3' direction, we found remnants of a second Tn3-like element specifying an active beta-lactamase. At their 5' extremities, the two incomplete copies of Tn3, which were in tandem orientation, were interrupted within the resolvase gene. We speculate that Tn3-related sequences have played a role in the process of selection and dissemination of the AAC(6')-4 gene, which specifies resistance to amikacin and related aminoglycosides.     

Domain dissection and characterization of the aminoglycoside resistance enzyme ANT(3″)-Ii/AAC(6′)-IId from Serratia marcescens

Aminoglycosides (AGs) are broad-spectrum antibiotics whose constant use and presence in growth environments has led bacteria to develop resistance mechanisms to aid in their survival. A common mechanism of resistance to AGs is their chemical modification (nucleotidylation, phosphorylation, or acetylation) by AG-modifying enzymes (AMEs). Through evolution, fusion of two AME-encoding genes has resulted in bifunctional enzymes with broader spectrum of activity. Serratia marcescens, a human enteropathogen, contains such a bifunctional enzyme, ANT(3″)-Ii/AAC(6′)-IId. To gain insight into the role, effect, and importance of the union of ANT(3″)-Ii and AAC(6′)-IId in this bifunctional enzyme, we separated the two domains and compared their activity to that of the full-length enzyme. We performed a thorough comparison of the substrate and cosubstrate profiles as well as kinetic characterization of the bifunctional ANT(3″)-Ii/AAC(6′)-IId and its individually expressed components.     

The use of aminoglycoside derivatives to study the mechanism of aminoglycoside 6'-N-acetyltransferase and the role of 6'-NH2 in antibacterial activity

Aminoglycoside antibiotics act by binding to 16S rRNA. Resistance to these antibiotics occurs via drug modifications by enzymes such as aminoglycoside 6'-N-acetyltransferases (AAC(6')s). We report here the regioselective and efficient synthesis of N-6'-acylated aminoglycosides and their use as probes to study AAC(6')-Ii and aminoglycoside-RNA complexes. Our results emphasize the central role of N-6' nucleophilicity for transformation by AAC(6')-Ii and the importance of hydrogen bonding between 6'-NH(2) and 16S rRNA for antibacterial activity.     

aac（6＇）-Ib-cr介导的喹喏酮类新耐药机制研究进展

AAC（6＇）-Ib是重要的氨基糖苷乙酰基团转移酶,其变异基因aac（6＇）-Ib-cr可同时作用于氨基糖苷类和氟喹诺酮类两类结构不同的抗生素,是引起细菌耐药性的一种重要作用机制。该文主要对aac（6＇）-Ib-cr介导的喹诺酮类新耐药机制相关研究进行综述。