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Virtual Screening of Peptide and Peptidomimetic Fragments Targeted to Inhibit Bacterial Dithiol Oxidase DsbA.

Duprez W, Bachu P, Stoermer MJ, Tay S, McMahon RM, Fairlie DP, Martin JL - PLoS ONE (2015)

Bottom Line: By targeting virulence rather than viability, development of resistance and side effects (through killing host native microbiota) might be minimized.Although only weakly potent relative to larger covalent peptide inhibitors that interact through the active site cysteine, these fragments offer new opportunities as templates to build non-covalent inhibitors.The results suggest that non-covalent peptidomimetics may need to interact with sites beyond the hydrophobic groove in order to produce potent DsbA inhibitors.

View Article: PubMed Central - PubMed

Affiliation: Division of Chemistry and Structural Biology, Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland, Australia.

ABSTRACT
Antibacterial drugs with novel scaffolds and new mechanisms of action are desperately needed to address the growing problem of antibiotic resistance. The periplasmic oxidative folding system in Gram-negative bacteria represents a possible target for anti-virulence antibacterials. By targeting virulence rather than viability, development of resistance and side effects (through killing host native microbiota) might be minimized. Here, we undertook the design of peptidomimetic inhibitors targeting the interaction between the two key enzymes of oxidative folding, DsbA and DsbB, with the ultimate goal of preventing virulence factor assembly. Structures of DsbB--or peptides--complexed with DsbA revealed key interactions with the DsbA active site cysteine, and with a hydrophobic groove adjacent to the active site. The present work aimed to discover peptidomimetics that target the hydrophobic groove to generate non-covalent DsbA inhibitors. The previously reported structure of a Proteus mirabilis DsbA active site cysteine mutant, in a non-covalent complex with the heptapeptide PWATCDS, was used as an in silico template for virtual screening of a peptidomimetic fragment library. The highest scoring fragment compound and nine derivatives were synthesized and evaluated for DsbA binding and inhibition. These experiments discovered peptidomimetic fragments with inhibitory activity at millimolar concentrations. Although only weakly potent relative to larger covalent peptide inhibitors that interact through the active site cysteine, these fragments offer new opportunities as templates to build non-covalent inhibitors. The results suggest that non-covalent peptidomimetics may need to interact with sites beyond the hydrophobic groove in order to produce potent DsbA inhibitors.

No MeSH data available.


Related in: MedlinePlus

Comparison of the docked designed peptidomimetic with the EcDsbA-EcDsbB and PmDsbA-PWATCDS crystal structures.Calculated electrostatic surfaces of the enzymes are shown, with acidic regions in red, basic regions in blue and non-polar (hydrophobic) regions in white. Electrostatics cut-offs used are +/- 7.5 keV. A. Detail of the EcDsbA complex with EcDsbB from the crystal structure (PDB code 2ZUP [37]) centred on the 97YPSPFATCDFMVR109 sequence of EcDsbB (in light blue) showing Phe101 (F101) binding in the EcDsbA hydrophobic groove (circled). B. Detail of the PmDsbAC30S:PWATCDS crystal structure (PDB code 4OD7) with PWATCDS in magenta. Residue Trp2 (W2) of the peptide binds in the PmDsbA hydrophobic groove (circled). C. Virtual screening identified compound 1 as a potential hit. Three optimal conformations of 1 are shown (in differing shades of green), in their predicted binding mode to the PmDsbAC30S hydrophobic groove. Potential hydrogen bonds between the morpholine moiety and DsbA Pro150, His32 and Asn162 are shown as yellow dashed lines.
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pone.0133805.g003: Comparison of the docked designed peptidomimetic with the EcDsbA-EcDsbB and PmDsbA-PWATCDS crystal structures.Calculated electrostatic surfaces of the enzymes are shown, with acidic regions in red, basic regions in blue and non-polar (hydrophobic) regions in white. Electrostatics cut-offs used are +/- 7.5 keV. A. Detail of the EcDsbA complex with EcDsbB from the crystal structure (PDB code 2ZUP [37]) centred on the 97YPSPFATCDFMVR109 sequence of EcDsbB (in light blue) showing Phe101 (F101) binding in the EcDsbA hydrophobic groove (circled). B. Detail of the PmDsbAC30S:PWATCDS crystal structure (PDB code 4OD7) with PWATCDS in magenta. Residue Trp2 (W2) of the peptide binds in the PmDsbA hydrophobic groove (circled). C. Virtual screening identified compound 1 as a potential hit. Three optimal conformations of 1 are shown (in differing shades of green), in their predicted binding mode to the PmDsbAC30S hydrophobic groove. Potential hydrogen bonds between the morpholine moiety and DsbA Pro150, His32 and Asn162 are shown as yellow dashed lines.

Mentions: The 1.6 Å resolution PmDsbAC30S-PWATCDS crystal structure PDB code 4OD7 [38], rather than the 3.7 Å resolution EcDsbB-EcDsbA crystal structure (PDB 2ZUP [40]), was used to analyse the hydrophobic groove interactions (Fig 3A). Virtual screening of a peptidomimetic library was performed by focusing on the interaction of Trp2 in heptapeptide PWATCDS with Tyr173 in PmDsbAC30S within the hydrophobic groove (Fig 3B). The compound with the best fit, hereafter named compound 1, was a tryptophan residue flanked by a C-terminal morpholine functional group and an N-terminal benzyl moiety (Fig 3C). Apart from the pi-stacking interaction between the Trp indole and Tyr173 of PmDsbAC30S, the ligand docking model showed the oxygen atom of the morpholine group at similar distances from the PmDsbAC30S Pro150 backbone amide, His32 imidazole ring amines, and the Asn162 side chain amide (≈ 4.5 Å). Compound 1 scored well in docking experiments performed using three different programs Goldscore, Chemscore and ChemPLP. Secondly, it had a good balance between aqueous solubility and "drug-like" properties (CLogP = 2.6, 6 rotatable bonds, 3 hydrogen bond acceptors, 3 hydrogen bond donors, MW = 406 Da, and is neutral at physiological pH). Compound 1 was an attractive first target for synthesis because it was predicted to extend its indole ring into the Trp-binding pocket, form hydrophobic interactions from its benzylamide group with the same region as the proline of the original peptide, and insert its morpholine ring into the groove.


Virtual Screening of Peptide and Peptidomimetic Fragments Targeted to Inhibit Bacterial Dithiol Oxidase DsbA.

Duprez W, Bachu P, Stoermer MJ, Tay S, McMahon RM, Fairlie DP, Martin JL - PLoS ONE (2015)

Comparison of the docked designed peptidomimetic with the EcDsbA-EcDsbB and PmDsbA-PWATCDS crystal structures.Calculated electrostatic surfaces of the enzymes are shown, with acidic regions in red, basic regions in blue and non-polar (hydrophobic) regions in white. Electrostatics cut-offs used are +/- 7.5 keV. A. Detail of the EcDsbA complex with EcDsbB from the crystal structure (PDB code 2ZUP [37]) centred on the 97YPSPFATCDFMVR109 sequence of EcDsbB (in light blue) showing Phe101 (F101) binding in the EcDsbA hydrophobic groove (circled). B. Detail of the PmDsbAC30S:PWATCDS crystal structure (PDB code 4OD7) with PWATCDS in magenta. Residue Trp2 (W2) of the peptide binds in the PmDsbA hydrophobic groove (circled). C. Virtual screening identified compound 1 as a potential hit. Three optimal conformations of 1 are shown (in differing shades of green), in their predicted binding mode to the PmDsbAC30S hydrophobic groove. Potential hydrogen bonds between the morpholine moiety and DsbA Pro150, His32 and Asn162 are shown as yellow dashed lines.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4520593&req=5

pone.0133805.g003: Comparison of the docked designed peptidomimetic with the EcDsbA-EcDsbB and PmDsbA-PWATCDS crystal structures.Calculated electrostatic surfaces of the enzymes are shown, with acidic regions in red, basic regions in blue and non-polar (hydrophobic) regions in white. Electrostatics cut-offs used are +/- 7.5 keV. A. Detail of the EcDsbA complex with EcDsbB from the crystal structure (PDB code 2ZUP [37]) centred on the 97YPSPFATCDFMVR109 sequence of EcDsbB (in light blue) showing Phe101 (F101) binding in the EcDsbA hydrophobic groove (circled). B. Detail of the PmDsbAC30S:PWATCDS crystal structure (PDB code 4OD7) with PWATCDS in magenta. Residue Trp2 (W2) of the peptide binds in the PmDsbA hydrophobic groove (circled). C. Virtual screening identified compound 1 as a potential hit. Three optimal conformations of 1 are shown (in differing shades of green), in their predicted binding mode to the PmDsbAC30S hydrophobic groove. Potential hydrogen bonds between the morpholine moiety and DsbA Pro150, His32 and Asn162 are shown as yellow dashed lines.
Mentions: The 1.6 Å resolution PmDsbAC30S-PWATCDS crystal structure PDB code 4OD7 [38], rather than the 3.7 Å resolution EcDsbB-EcDsbA crystal structure (PDB 2ZUP [40]), was used to analyse the hydrophobic groove interactions (Fig 3A). Virtual screening of a peptidomimetic library was performed by focusing on the interaction of Trp2 in heptapeptide PWATCDS with Tyr173 in PmDsbAC30S within the hydrophobic groove (Fig 3B). The compound with the best fit, hereafter named compound 1, was a tryptophan residue flanked by a C-terminal morpholine functional group and an N-terminal benzyl moiety (Fig 3C). Apart from the pi-stacking interaction between the Trp indole and Tyr173 of PmDsbAC30S, the ligand docking model showed the oxygen atom of the morpholine group at similar distances from the PmDsbAC30S Pro150 backbone amide, His32 imidazole ring amines, and the Asn162 side chain amide (≈ 4.5 Å). Compound 1 scored well in docking experiments performed using three different programs Goldscore, Chemscore and ChemPLP. Secondly, it had a good balance between aqueous solubility and "drug-like" properties (CLogP = 2.6, 6 rotatable bonds, 3 hydrogen bond acceptors, 3 hydrogen bond donors, MW = 406 Da, and is neutral at physiological pH). Compound 1 was an attractive first target for synthesis because it was predicted to extend its indole ring into the Trp-binding pocket, form hydrophobic interactions from its benzylamide group with the same region as the proline of the original peptide, and insert its morpholine ring into the groove.

Bottom Line: By targeting virulence rather than viability, development of resistance and side effects (through killing host native microbiota) might be minimized.Although only weakly potent relative to larger covalent peptide inhibitors that interact through the active site cysteine, these fragments offer new opportunities as templates to build non-covalent inhibitors.The results suggest that non-covalent peptidomimetics may need to interact with sites beyond the hydrophobic groove in order to produce potent DsbA inhibitors.

View Article: PubMed Central - PubMed

Affiliation: Division of Chemistry and Structural Biology, Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland, Australia.

ABSTRACT
Antibacterial drugs with novel scaffolds and new mechanisms of action are desperately needed to address the growing problem of antibiotic resistance. The periplasmic oxidative folding system in Gram-negative bacteria represents a possible target for anti-virulence antibacterials. By targeting virulence rather than viability, development of resistance and side effects (through killing host native microbiota) might be minimized. Here, we undertook the design of peptidomimetic inhibitors targeting the interaction between the two key enzymes of oxidative folding, DsbA and DsbB, with the ultimate goal of preventing virulence factor assembly. Structures of DsbB--or peptides--complexed with DsbA revealed key interactions with the DsbA active site cysteine, and with a hydrophobic groove adjacent to the active site. The present work aimed to discover peptidomimetics that target the hydrophobic groove to generate non-covalent DsbA inhibitors. The previously reported structure of a Proteus mirabilis DsbA active site cysteine mutant, in a non-covalent complex with the heptapeptide PWATCDS, was used as an in silico template for virtual screening of a peptidomimetic fragment library. The highest scoring fragment compound and nine derivatives were synthesized and evaluated for DsbA binding and inhibition. These experiments discovered peptidomimetic fragments with inhibitory activity at millimolar concentrations. Although only weakly potent relative to larger covalent peptide inhibitors that interact through the active site cysteine, these fragments offer new opportunities as templates to build non-covalent inhibitors. The results suggest that non-covalent peptidomimetics may need to interact with sites beyond the hydrophobic groove in order to produce potent DsbA inhibitors.

No MeSH data available.


Related in: MedlinePlus