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Crystal structure of New Delhi metallo-?-lactamase reveals molecular basis for antibiotic resistance.

ABSTRACT: ?-Lactams are the most commonly prescribed class of antibiotics and have had an enormous impact on human health. Thus, it is disquieting that an enzyme called New Delhi metallo-?-lactamase-1 (NDM-1) can confer Enterobacteriaceae with nearly complete resistance to all ?-lactam antibiotics including the carbapenams. We have determined the crystal structure of Klebsiella pneumoniae apo-NDM-1 to 2.1-Å resolution. From the structure, we see that NDM-1 has an expansive active site with a unique electrostatic profile, which we propose leads to a broader substrate specificity. In addition, NDM-1 undergoes important conformational changes upon substrate binding. These changes have not been previously observed in metallo-?-lactamase enzymes and may have a direct influence on substrate recognition and catalysis.

SUBMITTER: King D

PROVIDER: S-EPMC3190144 | biostudies-literature | 2011 Sep

REPOSITORIES: biostudies-literature

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Crystal structure of New Delhi metallo-β-lactamase reveals molecular basis for antibiotic resistance.

King Dustin D Strynadka Natalie N

Protein science : a publication of the Protein Society 20110802 9

β-Lactams are the most commonly prescribed class of antibiotics and have had an enormous impact on human health. Thus, it is disquieting that an enzyme called New Delhi metallo-β-lactamase-1 (NDM-1) can confer Enterobacteriaceae with nearly complete resistance to all β-lactam antibiotics including the carbapenams. We have determined the crystal structure of Klebsiella pneumoniae apo-NDM-1 to 2.1-Å resolution. From the structure, we see that NDM-1 has an expansive active site with a unique electr ...[more]

PMID: 21774017

Similar Datasets

Structure of New Delhi metallo-β-lactamase 1 (NDM-1).

Project description:Antibiotic resistance in bacterial pathogens poses a serious threat to human health and the metallo-β-lactamase (MBL) enzymes are responsible for much of this resistance. The recently identified New Delhi MBL 1 (NDM-1) is a novel member of this family that is capable of hydrolysing a wide variety of clinically important antibiotics. Here, the crystal structure of NDM-1 from Klebsiella pneumoniae is reported and its structure and active site are discussed in the context of other recently deposited coordinates of NDM-1.

| S-EPMC3212353 | biostudies-literature

Emergence of New Delhi metallo-β-lactamase, Austria.

Project description: Not available

| S-EPMC3204651 | biostudies-literature

Biochemical characterization of New Delhi metallo-?-lactamase variants reveals differences in protein stability.

Project description:Metallo-?-lactamase (MBL)-based resistance is a threat to the use of most ?-lactam antibiotics. Multiple variants of the New Delhi MBL (NDM) have recently been reported. Previous reports indicate that the substitutions affect NDM activity despite being located outside the active site. This study compares the biochemical properties of seven clinically reported NDM variants.NDM variants were generated by site-directed mutagenesis; recombinant proteins were purified to near homogeneity. Thermal stability and secondary structures of the variants were investigated using differential scanning fluorimetry and circular dichroism; kinetic parameters and MIC values were investigated for representative carbapenem, cephalosporin and penicillin substrates.The substitutions did not affect the overall folds of the NDM variants, within limits of detection; however, differences in thermal stabilities were observed. NDM-8 was the most stable variant with a melting temperature of 72°C compared with 60°C for NDM-1. In contrast to some previous studies, kcat/KM values were similar for carbapenem and penicillin substrates for NDM variants, but differences in kinetics were observed for cephalosporin substrates. Apparent substrate inhibition was observed with nitrocefin for variants containing the M154L substitution. In all cases, cefoxitin and ceftazidime were poorly hydrolysed with kcat/KM values <1 s(-1) ?M(-1).These results do not define major differences in the catalytic efficiencies of the studied NDM variants and carbapenem or penicillin substrates. Differences in the kinetics of cephalosporin hydrolysis were observed. The results do reveal that the clinically observed substitutions can make substantial differences in thermodynamic stability, suggesting that this may be a factor in MBL evolution.

| S-EPMC4291237 | biostudies-literature

New Delhi metallo-β-lactamase-producing Enterobacteriaceae, United States.

Project description:We characterized 9 New Delhi metallo-β-lactamase-producing Enterobacteriaceae (5 Klebsiella pneumoniae, 2 Escherichia coli, 1 Enterobacter cloacae, 1 Salmonella enterica serovar Senftenberg) isolates identified in the United States and cultured from 8 patients in 5 states during April 2009-March 2011. Isolates were resistant to β-lactams, fluoroquinolones, and aminoglycosides, demonstrated MICs ≤1 µg/mL of colistin and polymyxin, and yielded positive metallo-β-lactamase screening results. Eight isolates had blaNDM-1, and 1 isolate had a novel allele (blaNDM-6). All 8 patients had recently been in India or Pakistan, where 6 received inpatient health care. Plasmids carrying blaNDM frequently carried AmpC or extended spectrum β-lactamase genes. Two K. pneumoniae isolates and a K. pneumoniae isolate from Sweden shared incompatibility group A/C plasmids with indistinguishable restriction patterns and a common blaNDM fragment; all 3 were multilocus sequence type 14. Restriction profiles of the remaining New Delhi metallo-β-lactamase plasmids, including 2 from the same patient, were diverse.

| S-EPMC3713825 | biostudies-literature

Clinical Evolution of New Delhi Metallo-?-Lactamase (NDM) Optimizes Resistance under Zn(II) Deprivation.

Project description:Carbapenem-resistant Enterobacteriaceae (CRE) are rapidly spreading and taking a staggering toll on all health care systems, largely due to the dissemination of genes coding for potent carbapenemases. An important family of carbapenemases are the Zn(II)-dependent ?-lactamases, known as metallo-?-lactamases (MBLs). Among them, the New Delhi metallo-?-lactamase (NDM) has experienced the fastest and widest geographical spread. While other clinically important MBLs are soluble periplasmic enzymes, NDMs are lipoproteins anchored to the outer membrane in Gram-negative bacteria. This unique cellular localization endows NDMs with enhanced stability upon the Zn(II) starvation elicited by the immune system response at the sites of infection. Since the first report of NDM-1, new allelic variants (16 in total) have been identified in clinical isolates differing by a limited number of substitutions. Here, we show that these variants have evolved by accumulating mutations that enhance their stability or the Zn(II) binding affinity in vivo, overriding the most common evolutionary pressure acting on catalytic efficiency. We identified the ubiquitous substitution M154L as responsible for improving the Zn(II) binding capabilities of the NDM variants. These results also reveal that Zn(II) deprivation imposes a strict constraint on the evolution of this MBL, overriding the most common pressures acting on catalytic performance, and shed light on possible inhibitory strategies.

| S-EPMC5740384 | biostudies-literature

Pterostilbene restores carbapenem susceptibility in New Delhi metallo-?-lactamase-producing isolates by inhibiting the activity of New Delhi metallo-?-lactamases.

Project description:Background and purpose:Bacteria producing New Delhi metallo-?-lactamase-1 (NDM-1) are an increasing clinical threat. NDM-1 can inactivate almost all ?-lactams and is not sensitive to any existing ?-lactamase inhibitors. To identify effective inhibitors of the NDM-1 enzyme and clarify the mechanism of action, a "lead compound" for developing more potent NDM-1 inhibitors needs to be provided.Experimental approach:Natural compounds were tested by enzyme inhibition screening to find potential inhibitors. MIC assays, growth curve assays, and time-kill assays were conducted to evaluate the in vitro antibacterial activity of pterostilbene and the combination of pterostilbene and meropenem. A murine thigh model and a mouse pneumonia model were used to evaluate the in vivo efficacy of combined therapy. Molecular modelling and a mutational analysis were used to clarify the mechanism of action.Key results:Pterostilbene significantly inhibited NDM-1 hydrolysis activity in enzyme inhibition screening assays and effectively restored the effectiveness of meropenem in vitro with NDM-expressing isolates in antibacterial activity assays. In addition, the combined therapy effectively reduced the bacterial burden in a murine thigh model and protected mice from pneumonia caused by Klebsiella pneumoniae. By means of molecular dynamics simulation, we observed that pterostilbene localized to the catalytic pocket of NDM-1, hindering substrate binding to NDM-1 and reducing NDM-1 activity.Conclusions and implications:These findings indicated that pterostilbene combined with meropenem may offer a new safe and potential "lead compound" for the further development of NDM-1 inhibitors.

| S-EPMC6932935 | biostudies-literature

Dipicolinic Acid Derivatives as Inhibitors of New Delhi Metallo-?-lactamase-1.

Project description:The efficacy of ?-lactam antibiotics is threatened by the emergence and global spread of metallo-?-lactamase (MBL) mediated resistance, specifically New Delhi metallo-?-lactamase-1 (NDM-1). By utilization of fragment-based drug discovery (FBDD), a new class of inhibitors for NDM-1 and two related ?-lactamases, IMP-1 and VIM-2, was identified. On the basis of 2,6-dipicolinic acid (DPA), several libraries were synthesized for structure-activity relationship (SAR) analysis. Inhibitor 36 (IC50 = 80 nM) was identified to be highly selective for MBLs when compared to other Zn(II) metalloenzymes. While DPA displayed a propensity to chelate metal ions from NDM-1, 36 formed a stable NDM-1:Zn(II):inhibitor ternary complex, as demonstrated by 1H NMR, electron paramagnetic resonance (EPR) spectroscopy, equilibrium dialysis, intrinsic tryptophan fluorescence emission, and UV-vis spectroscopy. When coadministered with 36 (at concentrations nontoxic to mammalian cells), the minimum inhibitory concentrations (MICs) of imipenem against clinical isolates of Eschericia coli and Klebsiella pneumoniae harboring NDM-1 were reduced to susceptible levels.

| S-EPMC5599375 | biostudies-literature

Membrane anchoring stabilizes and favors secretion of New Delhi metallo-?-lactamase.

Project description:Carbapenems, 'last-resort' ?-lactam antibiotics, are inactivated by zinc-dependent metallo-?-lactamases (MBLs). The host innate immune response withholds nutrient metal ions from microbial pathogens by releasing metal-chelating proteins such as calprotectin. We show that metal sequestration is detrimental for the accumulation of MBLs in the bacterial periplasm, because those enzymes are readily degraded in their nonmetallated form. However, the New Delhi metallo-?-lactamase (NDM-1) can persist under conditions of metal depletion. NDM-1 is a lipidated protein that anchors to the outer membrane of Gram-negative bacteria. Membrane anchoring contributes to the unusual stability of NDM-1 and favors secretion of this enzyme in outer-membrane vesicles (OMVs). OMVs containing NDM-1 can protect nearby populations of bacteria from otherwise lethal antibiotic levels, and OMVs from clinical pathogens expressing NDM-1 can carry this MBL and the blaNDM gene. We show that protein export into OMVs can be targeted, providing possibilities of new antibacterial therapeutic strategies.

| S-EPMC4912412 | biostudies-literature

New Delhi Metallo-β-Lactamase-Producing Enterobacterales Bacteria, Switzerland, 2019-2020.

Project description:Carbapenemase-producing Enterobacterales (CPE) bacteria are a critical global health concern; New Delhi metallo-β-lactamase (NDM) enzymes account for >25% of all CPE found in Switzerland. We characterized NDM-positive CPE submitted to the Swiss National Reference Center for Emerging Antibiotic Resistance during a 2-year period (January 2019-December 2020) phenotypically and by using whole-genome sequencing. Most isolates were either Klebsiella pneumoniae (59/141) or Escherichia coli (52/141), and >50% were obtained from screening swabs. Among the 108 sequenced isolates, NDM-1 was the most prevalent variant, occurring in 56 isolates, mostly K. pneumoniae (34/56); the next most prevalent was NDM-5, which occurred in 49 isolates, mostly E. coli (40/49). Fourteen isolates coproduced a second carbapenemase, predominantly an OXA-48-like enzyme, and almost one third of isolates produced a 16S rRNA methylase conferring panresistance to aminoglycosides. We identified successful plasmids and global lineages as major factors contributing to the increasing prevalence of NDMs in Switzerland.

| S-EPMC8462332 | biostudies-literature

Structure, Genetics and Worldwide Spread of New Delhi Metallo-?-lactamase (NDM): a threat to public health.

Project description:The emergence of carbapenemase producing bacteria, especially New Delhi metallo-?-lactamase (NDM-1) and its variants, worldwide, has raised amajor public health concern. NDM-1 hydrolyzes a wide range of ?-lactam antibiotics, including carbapenems, which are the last resort of antibiotics for the treatment of infections caused by resistant strain of bacteria.In this review, we have discussed bla NDM-1variants, its genetic analysis including type of specific mutation, origin of country and spread among several type of bacterial species. Wide members of enterobacteriaceae, most commonly Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae, and gram-negative non-fermenters Pseudomonas spp. and Acinetobacter baumannii were found to carry these markers. Moreover, at least seventeen variants of bla NDM-type gene differing into one or two residues of amino acids at distinct positions have been reported so far among different species of bacteria from different countries. The genetic and structural studies of these variants are important to understand the mechanism of antibiotic hydrolysis as well as to design new molecules with inhibitory activity against antibiotics.This review provides a comprehensive view of structural differences among NDM-1 variants, which are a driving force behind their spread across the globe.

| S-EPMC5408368 | biostudies-literature

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