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SUPPLEMENTARY INFORMATIONDOI: 10.1038/NCHEM.2110

Supplementary Information

Rhodanine Hydrolysis Leads to Potent Thioenolate Mediated Metallo-β-Lactamase

Inhibition

Jürgen Brem,[a]+ Sander S. van Berkel,[a]+ WeiShen Aik,[a] Anna M. Rydzik,[a] Matthew B.

Avison,[b] Ilaria Pettinati,[a] Klaus-Daniel Umland,[a] Akane Kawamura,[a] James Spencer,[b]

Timothy D.W. Claridge,[a] Michael A. McDonough,[a] and Christopher J. Schofield[a]*

[a] Department of Chemistry, University of Oxford, 12 Mansfield Road, Oxford, OX1 3TA, United Kingdom [b] School of Cellular and Molecular Medicine, University of Bristol, Medical Sciences Building, Bristol, BS8 1TD, United Kingdom

E-mail: [email protected]

© 2014 Macmillan Publishers Limited. All rights reserved.



Content:

A) List of Figures and Schemes page S3

B) List of Tables page S11

C) Experimental section page S14

References page S19




A) List of Figures and Schemes

Supplementary Figure 1. Rhodanines and related compounds reported as PBP1, 2 and/or SBL3 inhibitors.

Supplementary Figure 2. View of the protein-ligand interactions in Chain A derived from a structure obtained by co-crystallization of ML302 with VIM-2. A) Protein-ligand interactions between VIM-2 and ML302F in Chain A; B) Protein-ligand interactions between VIM-2 and ML302 in Chain A. Protein-ligand interactions between VIM-2 and ML302M (ML302 and ML302F) depicted in monomer A using LIGPLOT.4 Hydrogen bonding interactions are shown as lines and ligand-protein hydrophobic contacts are shown as curved combs.




Supplementary Figure 3. Protein-ligand interactions in Chain B derived from a structure obtained by co-crystallization of ML302 with VIM-2. A) Protein-ligand interactions between VIM-2 and ML302F in chain B; B) Overlay of the protein-ligand interactions between VIM-2 and ML302F in Chains A and B. Protein-ligand interactions between VIM-2 and ML302M (ML302 and ML302F) depicted in monomer A using LIGPLOT.4 Hydrogen bonding interactions are shown as lines and ligand-protein hydrophobic contacts are shown as curved combs.

Supplementary Figure 4. Dihedral angles relating the plane of the 1,2,4-trichlorobenzene ring and the C-S bond; A’BCD: 106° and ABCD: 76° in Chain A, A’BCD: 103° and ABCD: 79° in Chain B for ML302F; A’BCD: 115° and ABCD: 69° for ML302.




Supplementary Figure 5. Proposed outline mechanism for ML302 hydrolysis leading to the thioenolate (ML302F). Studies on the chemical hydrolysis of rhodanines are reported,5 but enzyme catalyzed rhodanine hydrolysis is only reported for an imidase.6 MBL-catalyzed hydrolysis of thioesters is known (e.g. esters of thiomandelic acid),7 as well as the SBL and DD-transpeptidase (PBP) mediated (S)-selective hydrolysis of α-hydroxyalkyl esters.8 Thus, it is possible to envisage that rhodanines are hydrolysed by MBLs. However, although the hydrolysis lead to clear changes in the UV-VIS or 1H-NMR spectra, it was not possible to definitively assign MBL catalysed hydrolysis of ML302 due to the potency of inhibition.

Supplementary Figure 6. Time course analyses of hydrolysis of ML302 with or without added NDM-1 followed by 1H NMR (600 MHz). (A) in the presence of NDM-1; (B) in the absence of NDM-1.




Supplementary Figure 7. Crystallographically observed water displacement upon inhibitor binding to VIM-2. Views from crystal structures of A) apo-di-Zn(II)-VIM-2 (PDB ID: 1KO3),9 B) ML302:ML302F bound to VIM-2 (PDB ID: 4PVO) and C) ML302F bound to VIM-2 (PDB ID: 4PVT). Water molecules are shown as blue spheres; D) Close-up view of ML302F bound to VIM-2 (PDB ID: 4PVO). Water molecules are shown as green (apo-di-Zn(II)-VIM-2) and red (ML302:ML302F bound to VIM-2) spheres.




Supplementary Figure 8. Binding modes of the inhibitors thiomandelic acid and mercaptoacetic acid to MBLs. (A) Binding mode of (R)-thiomandelic acid to the Bacillus cereus metallo-β-lactamase (BcII, PDB ID: 2M5D) derived from an NMR structure of BcII complexed with thiomandelic acid.10 Note that, at least in the reported structure, thiomandelic acid carboxylate does not directly interact with the conserved Lys- and Asn-residues, which are known to be involved in substrate binding.11 B) Protein-ligand interactions between thiomandelic acid and BcII are depicted using LIGPLOT.4

Supplementary Figure 9. A) Binding mode of mercaptoacetic acid (MCR) to the MBL from Serratia marcescens (SMB-1, B3 subclass; PDB ID: 3VQZ) derived from a crystal structure of SMB-1 complexed with MCR.12 B) Protein-ligand interactions between mercaptoacetic acid and SMB-1 are depicted using LIGPLOT.4




Supplementary Figure 10. IC50 curves for ML302, ML302F and ML302M with a panel of MBLs.




Supplementary Figure 11. 19F NMR based ligand binding displacement assay for BcII. A) Titration of BcII in the solution of 4-(trifluoromethyl)thiophenol (3FCA, 50μM) yields a dose response curve for the binding of 3FCA to BcII. Data fitting gave an apparent Kd of 89.7±5.6 μM. B) and C) Displacement of 3FCA by ML302 and ML302F, respectively was observed for BcII, supporting the crystallographic observations. Note, an accurate Kd was possible to determine only for ML302F (μM) due to the poor solubility of ML302.




Supplementary Figure 12. 19F NMR time course analyses of NDM-1*-di-Zn(II)-ML302 or ML302F-complex. NDM-1*-di-Zn(II)-ML302 complex leads to a time-dependent decrease in the signal corresponding to the initial complex, due to the hydrolysis of ML302. The NDM-1*-di-Zn(II)-ML302F complex is stable over the same period of time.

Supplementary Figure 14. Cytotoxicity of ML302, ML302F and Staurosporine in HeLa cells.

-10 -8 -6 -4 -20

2

4

6

ML302FML302Staurosporin

Log concentration (M)

Abs

495




B) Tables

Supplementary Table 1. Crystallization conditions

Sample composition

Crystallization conditions

Vapour diffusion conditions

VIM-2:ML302 VIM-2 in crystallization buffer, TCEP (1 mM), ML302 (2.5 mM)

0.1 M magnesium formate, 25% PEG 3350

Sitting drop, protein-to-well ratio, 1:2, 293K

VIM-2:ML302F VIM-2 in crystallization buffer, TCEP (1 mM), ML302F (1 mM)

0.1 M magnesium formate, 23% PEG 3350


BcII:ML302F BcII in crystallization buffer, TCEP (1 mM), ML302F (5 mM)

0.2 M ammonium sulfate, 0.1 M Bis-Tris pH = 5.5, 25% PEG 3350





Supplementary Table 2. Data Collection and Refinement Statistics

§ Rmerge =∑j∑h| Ihj – <Ih>| /∑j∑h <Ih>×100

* Rwork =∑||Fobs| – |Fcalc||/|Fobs|×100

† Rfree, based on 2.9% (4PVO), 7.4% (4PVT) and 9.69 % (4TYT) of the total reflections ‡ outermost shell

Structure VIM2-ML302 VIM2-ML302F BcII-ML302F

PDB ID 4PVO 4PVT 4TYT

Radiation Source Diamond I04 Rotating anode Rotating anode

Resolution Range (Å) 50.0 – 1.48 (1.53 – 1.48) ‡

50.0 – 2.00 (2.07 – 2.00) ‡

28.0 – 1.80 (1.86 – 1.80) ‡

Space Group C 1 2 1 C 1 2 1 C 1 2 1

Unit Cell Dimensions

a, b, c (Å) 104.57, 79.24, 67.97 102.39, 78.91, 67.74 53.08, 61.36, 69.42

α, β, γ (˚) 90.00, 131.57, 90.00 90.00, 130.32, 90.00 90.00, 93.00, 90.00

Total Number of Reflections Observed

339434 97523 57530

Number of Unique Reflections

68140 27185 20706

Redundancy 5.0 (4.1) ‡ 3.6 (3.4) ‡ 2.9 (1.7) ‡

Completeness (%) 99.0 (93.9) ‡ 98.0 (96.4) ‡ 99.7 (99.1) ‡

I/σ(I) 13.0 (2.4) ‡ 6.8 (2.5) ‡ 7.86 (1.56) ‡

§Rmerge 0.136 (0.568) ‡ 0.175 (0.521) ‡ 0.108 (0.366) ‡

*Rwork (%) 14.08 16.58 15.00

† Rfree (%) 16.46 21.89 20.13

RMS Deviation from Ideality

bonds 0.0130 Å 0.0098 Å 0.006 Å

angles 1.518° 1.209° 0.986°

Average B Factor (Å2) 16.76 21.74 24.23

Wilson B Factor (Å2) 10.65 17.32 18.06

Number of Water Molecules

536 351 264




Supplementary Table 3. Tests for inhibition by ML302 and ML302F of a panel of known rhodanine targets2, 3, 13 and human enzymes under standard conditions to investigate the selectivity of ML302 and ML302F.

Entry Enzyme ML302 ML302F Control

RA [%] RA [%]a

1 TEM-1 98.6±7.1/N.I. 99.6±3.1/N.I. Clavulanic acid

2 PBP5 (dacA) from E. coli 104.6±5.7 99.2±6.4/N.I. Piperacillin

3 PBP6 (dacC) from E. coli 97.7±3.8/N.I. 114±19.6/N.I. Piperacillin

4 hHAT 93±15/N.I. 85±9/N.I. Anacardic acid

5 hHDAC 108±3/N.I. 116±7/N.I. Trichostatin

6 hACE-2 73±9 99±4/N.I. EDTA

7 hGlyoxylase II 98±3/N.I. 95±6/N.I. EDTA

N.I. – not inhibited; RA ˂ 1% at 100 μM.




C) Experimental section

Material and Methods

Chemicals were from commonly used suppliers and were used without further purification. Solvents (including dry solvents) for chemical transformations, work-up and chromatography were from Sigma-Aldrich (Dorset, UK) at HPLC grade, and used without further distillation. Silica gel 60 F254 analytical thin layer chromatography (TLC) plates were from Merck (Darmstadt, Germany) and visualized under UV light and/or with potassium permanganate stain. Chromatographic purifications were performed using Merck Geduran 60 silica (40-63 μm) or prepacked SNAP columns using a Biotage SP1 Purification system (Uppsala, Sweden). Microwave assisted reactions were performed using a Biotage InitiatorTM microwave synthesizer in sealed vials. Deuterated solvents were obtained from Chambridge Isotopes and Apollo Scientific Ltd. All 1H and 13C NMR spectra were recorded using a Bruker Avance 400 MHz spectrometer. All chemical shifts are given in ppm relative to the solvent peak,14 and coupling constants (J) are reported in Hz to the closed 0.5. High Resolution (HR) mass spectrometry data (m/z) were obtained from a Bruker MicroTOF instrument using an ESI source and Time of Flight (TOF) analyzer. Low Resolution (LR) mass spectrometry data (m/z) were obtained from a Waters LCT Premier instrument using an ESI source and Time of Flight (TOF) analyzer. Melting points were obtained using a Stuart SMP-40 automatic melting point apparatus.

Synthesis15

(Z)-2-(4-Oxo-2-thioxo-5-(2,3,6-trichlorobenzylidene)thiazolidin-3-yl)acetic acid (1)15

A 20-mL microwave vial was charged with 2,3,6-trichlorobenzaldehdye (1.05 g, 5.0 mmol) and rhodanine-3-acetic acid (955 mg, 5.0 mmol). Ethanol (10 mL) was added followed by piperidine (4 drops). The vial was sealed and submitted to microwave irradiation at 120 °C for 90 min. The solvent was removed and the crude product purified by column chromatography (CH2Cl2/MeOH 9:1). The desired product was obtained as light yellow solid (1.35 g, 71%). TLC (CH2Cl2/MeOH 9:1) Rf = 0.55. 1H NMR (400 MHz CDCl3) δ = 8.50- 7.00 (br. s, COOH, 1H), 7.76 (s, 1H), 7.47 - 7.52 (m, 1H), 7.36 (d, J = 9.0 Hz, 1H), 4.93 (s, 2H) ppm. 13C NMR (101 MHz CDCl3) δ = 191.9, 170.9, 165.2, 132.6, 132.3, 132.2, 131.8, 131.31, 131.29, 128.7, 128.1, 44.1 ppm. LRMS m/z calcd. for C12H5Cl3NO3S2 [M-H] = 379.88, found [M-H] = 379.8.




N-(4-Methylpiperazin-1-yl)-2-(4-oxo-2-thioxo-5-(2,3,6-trichlorobenzylidene)thiazolidin-3-yl)acetamide (E/Z mixture (1:3)) (ML302)15

A solution of 1 (381 mg, 1.0 mmol) and Bis(2-oxo-3-oxazolidinyl)phosphonic chloride (BOP-Cl, 350 mg, 1.2 mmol, 1.2 equiv) in CH2Cl2 (10 mL) was cooled to 0 °C using an ice/water bath under an inert atmosphere. Freshly distilled triethylamine (Et3N, 168 μL, 1.2 mmol, 1.2 equiv) was added and the solution was allowed to stir at 0 °C for 30 min. 1-Amino-4-methyl-piperazine (145 μL, 1.2 mmol, 1.2 equiv,) was added and the temperature was raised to r.t. after which the reaction was stirred overnight. The crude product, obtained by removal of dichloromethane in vacuo, was than purified by flash chromatography (CH2Cl2/MeOH 9:1) to give compound ML302 as a non-separated mixture of E/Z isomers in a 1:3 ratio (373 mg, 78%) as a yellow solid. TLC (CH2Cl2/MeOH 9:1) Rf = 0.35. 1H NMR (400 MHz, CDCl3) δ (assigned for the major isomer) = 7.72 (s, 1H), 7.47 (dd, J = 8.5, 0.5 Hz, 1H), 7.34 (d, J = 9.0 Hz, 2H), 6.90 (br. s., 1H), 5.05 (s, 2H), 2.95 - 3.20 (m, 2H), 2.65 - 2.90 (m, 4H), 2.33 (s, 3H) ppm. 13C NMR (101 MHz CDCl3) δ (assigned for the major isomer) = 192.9, 166.9, 166.0, 133.1, 132.5, 132.08, 132.05, 128.9, 127.9, 56.0, 54.2 (4C), 45.4, 44.9 ppm. LRMS m/z calcd. for C17H18Cl3N4O2S2 [M+H] = 478.99, found [M+H] = 479.0. Mp = 182–185 °C.

2-Mercapto-3-(2,3,6-trichlorophenyl)acrylic acid (E/Z mixture (1:4) (ML302F)

Compound ML302 (240 mg, 0.5 mmol) was dissolved in 1M NaOH (4 mL) upon which a colour change to deep red was observed. The clear reaction mixture was warmed to 60 °C, then stirred for 45 min (the reaction colour changes from deep red to yellow) when consumption of starting material was observed. The reaction mixture was cooled to room temperature and subsequently carefully acidified with 1M HCl (5 mL). A white precipitate was formed and collected by centrifugation (4000g/10 min). After removal of the liquid phase the white solid was resuspended in H2O (2 mL) and again collected by centrifugation. This process was repeated twice. The obtained solid was dried under high vacuum overnight. The product was obtained as an inseparable mixture of E/Z isomers in a 1:4 ratio as an off-white solid (40 mg, 28%). TLC (CH2Cl2/MeOH 4:1) Rf = 0.67. 1H NMR (400 MHz, CDCl3) δ (assigned for the major isomer) = 7.69 (s, 1H), 7.45 (dd, J = 8.5, 1.0 Hz, 1H), 7.33 - 7.37




(m, 1H), 3.89 (br. s., 1H) ppm. 13C-NMR (200 MHz, CDCl3) δ = 168.1, 134.7, 132.8, 132.4, 132.1, 132.1, 130.7, 130.6, 128.7 ppm. LRMS m/z calcd. for C9H4Cl3O2S [M-H] = 280.90, found [M-H] =280.8.

Sodium 2-mercapto-3-(2,3,6-trichlorophenyl)acrylate (ML302F-Na) was obtained by dissolving ML302F (28,2 mg, 0.1 mmol) in 0.1M NaHCO3 (1 mL). The resulting clear solution was subsequently lyophilized to generate the desired sodium salt as an off-white solid (31 mg, quant). LRMS m/z calcd. for C9H4Cl3NaO2S [M+Na] = 303.89, found [M-H] = 280.8.

Protein production and purification

Recombinant forms of NDM-1, NDM-1 M67C, VIM-1, VIM-2, SPM-1, IMP-1 and BcII MBLs were produced in Escherichia coli as described.16 Purified VIM-2 protein was dialysed into fresh crystallization buffer (50mM HEPES pH 7.5, 150 mM NaCl and 1 µg ZnCl2, concentrated to 7.5 mg/mL and used for crystallographic analyses.

Crystallography

Crystals of VIM-2:ML302, VIM-2:ML302F and BcII:ML302F were grown using the conditions stated in Table S1. The crystals were cryoprotected using well solution diluted with 25% glycerol before being flash cooled in liquid nitrogen. Data for VIM-2:ML302 were collected at Diamond Light Source synchrotron beamline I04 and data for VIM-2:ML302F and BcII:ML302F were collected using a Rigaku FRE+ Superbright diffractometer. All data sets were collected at 100K. All data were indexed, integrated and scaled using HKL-3000.17 The VIM-2:ML302 structure was solved by molecular replacement using Phaser18 using the VIM-2 structure (PDB ID: 1KO3)9 as a search model. The VIM-2:ML302 structure was then used as a search model for the VIM-2:ML302F structure. The BcII:ML302F structure was solved by molecular replacement using Phaser18 and our unreleased BcII structure (PDB ID: 4C09) as a search model. The structures were then refined using PHENIX19 and COOT20 until Rwork and Rfree no longer converged. Data collection and refinement statistics are given in Table S2.




Kinetic analyses

Kinetic and inhibition analyses and UV-VIS studies16 against bacterial MBLs were performed as described.21

NDM-1 M67C labelling

NDM-1 M67C labelling with 3-bromo-1,1,1-trifluoroacetone (BFA) was performed via cysteine alkylation as described.22

NMR experiments 19F NMR experiments

19F NMR samples were prepared as follows: 0.09 mM labelled NDM-1/SPM-1/ 0.05 mM unlabelled BcII in 50 mM Tris/HCl buffer pH 7.5 (90% H2O, 10% D2O, 200 mM NaCl) was treated with 0.2 mM of desired compound. 19F NMR experiments were recorded using a Bruker AVIII 600 with BB-F/1H Prodigy N2 cryoprobe operating at 298K using 5 mm diameter NMR tubes (Norell). 19F 90o pulse lengths were 15 μs and spectra were typically obtained using 256 scans and a recovery delay of 2s. Data were processed with 1 Hz Lorentzian line broadening using TopSpin 3.1 software (Bruker) and were referenced to the internal TFA standard (0.05 mM, -75.45 ppm).

1H NMR experiments

Samples for ML302 hydrolysis were prepared as follows: 0.2 mM ML302 was dissolved in 50 mM Tris-d11/HCl buffer (90% H2O, 10% D2O) pH 7.5. Spectra were recorded in the absence or presence of NDM-1 (1 µM).

1H NMR time-courses were recorded either on Bruker AVIII 700 with 1H/13C/15N TCI cryoprobe or Bruker AVIII 600 with BB-F/1H Prodigy N2 cryoprobe with a typical 256 scans (AVIII 600) or 128 scans (AVIII 700) and 90o pulse lengths were of 12 μs. Data were processed with 1 Hz (AVIII 600) or 0.3 Hz (AVIII 700) Lorentzian line broadening using TopSpin 3.1 software (Bruker).

TEM-1 and Penicillin binding protein productions

TEM-1, PBP5 (dacA) from E. coli, PBP6 (dacC) from E. coli and PBP3 from P. aeruginosa were produced as reported.23




Inhibition studies for penicillin binding proteins

The inhibition analyses were performed using the method described for PBP-3.24 In brief:

ML302 and ML302F were initially screened at a single concentration (100 μM) in duplicate.

Inhibition of the PBP acylation rate by the chromogenic β-lactam Nitrocefin was determined

by pre-incubation of the appropriate amount of enzyme (dacA 2µM and dacC 6.8 µM,

respectively) and inhibitor in the assay buffer (50 mM HEPES-NaOH buffer (pH 7.2)

supplemented with 200mM NaCl and 0.01 % Triton 100) for 10 min. at r.t. prior to the

initiation of the assay by the addition of nitrocefin (100 µM final concentration). The

hydrolysis of Nitrocefin was monitored by following the variation in absorbance at 492 nm

using 96 well flat bottom plates (UV-STAR Microplate (655801) or Greiner Bio-One). The

experiments were performed using a PHERAstar FS – BMG Labtech microplate reader

(using path length correction) at r.t. (24-25°C). All enzymes and inhibitors were prepared in

the assay buffer.

Inhibition studies TEM-1

The inhibition analysis for TEM-1 was performed using the modified method as previously

described for MBLs.21 In brief: ML302 and ML302F were screened at a single concentration

(100 μM) in duplicate. Inhibition was tested by pre-incubation of the appropriate amount of

TEM-1 (1 nM final concentration) and ML302 or ML302F in the assay buffer (50 mM

HEPES-NaOH buffer (pH 7.2) supplemented with 0.01 % Triton 100) for 10 min. at r.t. prior

to the initiation of the assay by the addition of FC5 ((6R,7R)-8-Oxo-3-(((2-oxo-2H-chromen-

7-yl)oxy)methyl)-7-(2-phenylacetamido)-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic

acid 5,5-dioxide)21 (10 µM final concentration). The hydrolysis of FC5 was monitored by

following the variation on fluorescence reading (380nm – excitation and 460nm - emission)

using 384 well flat bottom plates (Greiner Bio-One). The experiments were performed on a

PHERAstar FS – BMG Labtech microplate reader at r.t. (24-25°C). All enzymes and

inhibitors were prepared in the assay buffer.

Inhibition studies for hHAT, HDAC, hACE-2 and hHAGH

The inhibition analyses for human histone acetyltransferases (HAT), HDAC hHDAC (human histone deacetylase), hACE-2 (Angiotensin converting enzyme 2) and hHAGH (Hydroxyacylglutathione hydrolase, human Glyoxylase II) were performed using the commercially available HAT Assay Kit (Active Motif, catalogue No. 56100), hHDAC (Active Motif, catalogue No. 56200), hACE-2 (ACE-2 (R&D Systems) and hHAGH ((R&D Systems).




References

1. Fedarovich, A. et al. High-Throughput Screening for Novel Inhibitors of Neisseria gonorrhoeae Penicillin-Binding Protein 2. PLoS ONE, 7, e44918, (2012).

2. Zervosen, A. et al. Interactions between Penicillin-Binding Proteins (PBPs) and Two Novel Classes of PBP Inhibitors, Arylalkylidene Rhodanines and Arylalkylidene Iminothiazolidin-4-ones. Antimicrob. Agents Chem. 48, 961-969 (2004).

3. Grant, E.B. et al. The synthesis and SAR of rhodanines as novel class C β-lactamase inhibitors. Bioorg.Med. Chem. Lett. 10, 2179-2182 (2000).

4. Wallace, A.C., Laskowski, R.A. & Thornton, J.M. LIGPLOT: a program to generate schematic diagrams of protein-ligand interactions. Protein Eng. 8, 127-34 (1995).

5. Campaigne, E. & Cline, R.E. Preparation and Absorption Spectra of Some β-Aryl-α-Mercaptoacrylic Acids and Related Disulfides. J. Org. Chem. 21, 32-38 (1956).

6. Ogawa, J., Soong, C.L., Honda, M. & Shimizu, S. Imidase, a dihydropyrimidinase-like enzyme involved in the metabolism of cyclic imides. Eur. J. Biochem. 243, 322-327 (1997).

7. Mollard, C. et al. Thiomandelic acid, a broad spectrum inhibitor of zinc β-lactamases. Kinetic and spectroscopic studies. J. Biol. Chem. 276, 45015-45023 (2001).

8. Pelto, R.B. & Pratt, R.F. Kinetics and stereochemistry of hydrolysis of an N-(phenylacetyl)-alpha-hydroxyglycine ester catalyzed by serine β-lactamases and DD-peptidases. Org. Biomol. Chem. 10, 7356-62 (2012).

9. Garcia-Saez, I., Docquier, J.D., Rossolini, G.M. & Dideberg, O. The Three-Dimensional Structure of VIM-2, a Zn-β-Lactamase from Pseudomonas aeruginosa in Its Reduced and Oxidised Form. J. Mol. Biol. 375, 604-611 (2008).

10. Karsisiotis, A.I., Damblon, C.F. & Roberts, G.C. Solution structures of the Bacillus cereus metallo-β-lactamase BcII and its complex with the broad spectrum inhibitor R-thiomandelic acid. Biochem J. 456, 397-407 (2013).

11. Fast, W. & Sutton, L.D. Metallo-β-lactamase: inhibitors and reporter substrates. Biochim. Biophys. Acta 1834, 1648-59 (2013).

12. Wachino, J. et al. Structural insights into the subclass B3 metallo-β-lactamase SMB-1 and the mode of inhibition by the common metallo-β-lactamase inhibitor mercaptoacetate. Antimicrob Agents Chemother 57, 101-9 (2013).

13. Furdas, S.D., Shekfeh, S., Kannan, S., Sippl, W., Jung, M. Rhodanine carboxylic acids as novel inhibitors of histone acetyltransferases. MedChemComm 3, 305-311 (2012).

14. Gottlieb, H.E., Kotlyar, V. & Nudelman, A. NMR Chemical Shifts of Common Laboratory Solvents as Trace Impurities. J. Org. Chem. 62, 7512-7515 (1997).

15. Spicer, T. et al. ML302: A Novel Beta-lactamase (BLA) Inhibitor. in Probe Reports from the NIH Molecular Libraries Program (National Center for Biotechnology Information (US), Bethesda (MD), 2010).

16. Makena, A. et al. Chromophore-linked substrate (CLS405): probing metallo-β-lactamase activity and inhibition. ChemMedChem 8, 1923-9 (2013).

17. Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Vol. 276 307-326 (Elsevier, 1997).

18. McCoy, A.J. et al. Phaser crystallographic software. J. Appl. Crystallogr. 40, 658-674 (2007).

19. Adams, P.D. et al. PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr. D Biol. Crystallogr. 66, 213-21 (2010).

20. Emsley, P., Lohkamp, B., Scott, W.G. & Cowtan, K. Features and development of Coot. Acta Crystallogr. D Biol. Crystallogr. 66, 486-501 (2010).

21. van Berkel, S.S. et al. Assay Platform for Clinically Relevant Metallo-β-lactamases. J. Med. Chem. 56, 6945-6953 (2013).




22. Rydzik, A.M. et al. Monitoring Conformational Changes in the NDM-1 Metallo-β-lactamase by 19F NMR Spectroscopy. Angew. Chem. Int. Ed. 53, 3129-33 (2014).

23. Inglis, S.R. et al. A boronic-acid-based probe for fluorescence polarization assays with penicillin binding proteins and beta-lactamases. Anal. Biochem., 420, 41-47 (2012).

24. van Berkel, S.S. et al. Binding of (5 S)-penicilloic acid to penicillin binding protein 3. ACS Chem. Biol. 8, 2112-2116 (2013).


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