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Structure-assisted discovery of an aminothiazole derivative as a lead molecule for inhibition of bacterial fatty-acid synthesis.

Pappenberger G, Schulz-Gasch T, Kusznir E, Müller F, Hennig M - Acta Crystallogr. D Biol. Crystallogr. (2007)

Bottom Line: The active site is accessible through an open conformation of the Phe392 side chain and no conformational changes are induced at the active site upon ligand binding.This represents a novel binding mode that differs from thiolactomycin or cerulenin interaction.The structural information on the protein-ligand interaction offers strategies for further optimization of this low-molecular-weight compound.

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Affiliation: F. Hoffmann-La Roche Ltd, Pharma Research Discovery, CH-4070 Basel, Switzerland.

ABSTRACT
Fatty-acid synthesis in bacteria is of great interest as a target for the discovery of antibacterial compounds. The addition of a new acetyl moiety to the growing fatty-acid chain, an essential step in this process, is catalyzed by beta-ketoacyl-ACP synthase (KAS). It is inhibited by natural antibiotics such as cerulenin and thiolactomycin; however, these lack the requirements for optimal drug development. Structure-based biophysical screening revealed a novel synthetic small molecule, 2-phenylamino-4-methyl-5-acetylthiazole, that binds to Escherichia coli KAS I with a binding constant of 25 microM as determined by fluorescence titration. A 1.35 A crystal structure of its complex with its target reveals noncovalent interactions with the active-site Cys163 and hydrophobic residues of the fatty-acid binding pocket. The active site is accessible through an open conformation of the Phe392 side chain and no conformational changes are induced at the active site upon ligand binding. This represents a novel binding mode that differs from thiolactomycin or cerulenin interaction. The structural information on the protein-ligand interaction offers strategies for further optimization of this low-molecular-weight compound.

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Binding mode of the aminothiazole ligand to E. coli KAS I protein. (a) Electron density of the aminothiazole ligand (green) bound to the active site of E. coli KAS I protein (grey). The 2F                  o − F                  c OMIT map (contoured at 1σ) is shown in black; the F                  o − F                  c OMIT map (contoured at 2.5σ) is shown in orange. The OMIT map in the absence of inhibitor was calculated using the final refined model. (b) Superposition of the binding modes for the KAS I ligands thiolactomycin (cyan), cerulenin (gold) and aminothiazole (green). For the representation of the protein, the structure with bound aminothiazole ligand was used. The protein is shown in surface representation, coloured according to atom type (C, grey; N, blue; O, red; S, yellow). Monomer B of the dimer has been removed, as well as side chains Met204 and Val270 of monomer A, to enable insight into the binding pocket. With an intact binding site, only the acetyl moiety of the aminothiazole ligand is exposed to solvent. The isoprenoid tail of thiolactomycin is buried in a hydrophobic pocket and is not visible in this representation. The structure and binding mode of cerulenin was taken from PDB entry 1fj8 (Price et al., 2001 ▶).
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fig3: Binding mode of the aminothiazole ligand to E. coli KAS I protein. (a) Electron density of the aminothiazole ligand (green) bound to the active site of E. coli KAS I protein (grey). The 2F o − F c OMIT map (contoured at 1σ) is shown in black; the F o − F c OMIT map (contoured at 2.5σ) is shown in orange. The OMIT map in the absence of inhibitor was calculated using the final refined model. (b) Superposition of the binding modes for the KAS I ligands thiolactomycin (cyan), cerulenin (gold) and aminothiazole (green). For the representation of the protein, the structure with bound aminothiazole ligand was used. The protein is shown in surface representation, coloured according to atom type (C, grey; N, blue; O, red; S, yellow). Monomer B of the dimer has been removed, as well as side chains Met204 and Val270 of monomer A, to enable insight into the binding pocket. With an intact binding site, only the acetyl moiety of the aminothiazole ligand is exposed to solvent. The isoprenoid tail of thiolactomycin is buried in a hydrophobic pocket and is not visible in this representation. The structure and binding mode of cerulenin was taken from PDB entry 1fj8 (Price et al., 2001 ▶).

Mentions: Crystal soaking with the aminothiazole compound in PEG 8000 in the presence of TCEP yielded a 1.35 Å data set (Table 1 ▶) that revealed strong electron density for the bound ligand at the active site in the F o − F c map (Fig. 3 ▶ a). This data set provides the highest resolution structure of E. coli KAS I obtained to date, enabling unambiguous assignment of the ligand-binding mode, of protein side-chain conformations (such as Phe392) and the positioning of water molecules (such as HOH2433) around the active site to aid in interpretation of the structural information.


Structure-assisted discovery of an aminothiazole derivative as a lead molecule for inhibition of bacterial fatty-acid synthesis.

Pappenberger G, Schulz-Gasch T, Kusznir E, Müller F, Hennig M - Acta Crystallogr. D Biol. Crystallogr. (2007)

Binding mode of the aminothiazole ligand to E. coli KAS I protein. (a) Electron density of the aminothiazole ligand (green) bound to the active site of E. coli KAS I protein (grey). The 2F                  o − F                  c OMIT map (contoured at 1σ) is shown in black; the F                  o − F                  c OMIT map (contoured at 2.5σ) is shown in orange. The OMIT map in the absence of inhibitor was calculated using the final refined model. (b) Superposition of the binding modes for the KAS I ligands thiolactomycin (cyan), cerulenin (gold) and aminothiazole (green). For the representation of the protein, the structure with bound aminothiazole ligand was used. The protein is shown in surface representation, coloured according to atom type (C, grey; N, blue; O, red; S, yellow). Monomer B of the dimer has been removed, as well as side chains Met204 and Val270 of monomer A, to enable insight into the binding pocket. With an intact binding site, only the acetyl moiety of the aminothiazole ligand is exposed to solvent. The isoprenoid tail of thiolactomycin is buried in a hydrophobic pocket and is not visible in this representation. The structure and binding mode of cerulenin was taken from PDB entry 1fj8 (Price et al., 2001 ▶).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2483479&req=5

fig3: Binding mode of the aminothiazole ligand to E. coli KAS I protein. (a) Electron density of the aminothiazole ligand (green) bound to the active site of E. coli KAS I protein (grey). The 2F o − F c OMIT map (contoured at 1σ) is shown in black; the F o − F c OMIT map (contoured at 2.5σ) is shown in orange. The OMIT map in the absence of inhibitor was calculated using the final refined model. (b) Superposition of the binding modes for the KAS I ligands thiolactomycin (cyan), cerulenin (gold) and aminothiazole (green). For the representation of the protein, the structure with bound aminothiazole ligand was used. The protein is shown in surface representation, coloured according to atom type (C, grey; N, blue; O, red; S, yellow). Monomer B of the dimer has been removed, as well as side chains Met204 and Val270 of monomer A, to enable insight into the binding pocket. With an intact binding site, only the acetyl moiety of the aminothiazole ligand is exposed to solvent. The isoprenoid tail of thiolactomycin is buried in a hydrophobic pocket and is not visible in this representation. The structure and binding mode of cerulenin was taken from PDB entry 1fj8 (Price et al., 2001 ▶).
Mentions: Crystal soaking with the aminothiazole compound in PEG 8000 in the presence of TCEP yielded a 1.35 Å data set (Table 1 ▶) that revealed strong electron density for the bound ligand at the active site in the F o − F c map (Fig. 3 ▶ a). This data set provides the highest resolution structure of E. coli KAS I obtained to date, enabling unambiguous assignment of the ligand-binding mode, of protein side-chain conformations (such as Phe392) and the positioning of water molecules (such as HOH2433) around the active site to aid in interpretation of the structural information.

Bottom Line: The active site is accessible through an open conformation of the Phe392 side chain and no conformational changes are induced at the active site upon ligand binding.This represents a novel binding mode that differs from thiolactomycin or cerulenin interaction.The structural information on the protein-ligand interaction offers strategies for further optimization of this low-molecular-weight compound.

View Article: PubMed Central - HTML - PubMed

Affiliation: F. Hoffmann-La Roche Ltd, Pharma Research Discovery, CH-4070 Basel, Switzerland.

ABSTRACT
Fatty-acid synthesis in bacteria is of great interest as a target for the discovery of antibacterial compounds. The addition of a new acetyl moiety to the growing fatty-acid chain, an essential step in this process, is catalyzed by beta-ketoacyl-ACP synthase (KAS). It is inhibited by natural antibiotics such as cerulenin and thiolactomycin; however, these lack the requirements for optimal drug development. Structure-based biophysical screening revealed a novel synthetic small molecule, 2-phenylamino-4-methyl-5-acetylthiazole, that binds to Escherichia coli KAS I with a binding constant of 25 microM as determined by fluorescence titration. A 1.35 A crystal structure of its complex with its target reveals noncovalent interactions with the active-site Cys163 and hydrophobic residues of the fatty-acid binding pocket. The active site is accessible through an open conformation of the Phe392 side chain and no conformational changes are induced at the active site upon ligand binding. This represents a novel binding mode that differs from thiolactomycin or cerulenin interaction. The structural information on the protein-ligand interaction offers strategies for further optimization of this low-molecular-weight compound.

Show MeSH
Related in: MedlinePlus