Dataset Information

Antagonism of a zinc metalloprotease using a unique metal-chelating scaffold: tropolones as inhibitors of P. aeruginosa elastase.

ABSTRACT: Tropolone emerged from the screening of a chelator fragment library (CFL) as an inhibitor of the Zn(2+)-dependent virulence factor, Pseudomonas aeruginosa elastase (LasB). Based on this initial hit, a series of substituted tropolone-based LasB inhibitors was prepared, and a compound displaying potent activity in vitro and in a bacterial swarming assay was identified. Importantly, this inhibitor was found to be specific for LasB over other metalloenzymes, validating the usage of tropolone as a viable scaffold for identifying first-in-class LasB inhibitors.

SUBMITTER: Fullagar JL

PROVIDER: S-EPMC3618488 | biostudies-literature | 2013 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Antagonism of a zinc metalloprotease using a unique metal-chelating scaffold: tropolones as inhibitors of P. aeruginosa elastase.

Fullagar Jessica L JL Garner Amanda L AL Struss Anjali K AK Day Joshua A JA Martin David P DP Yu Jing J Cai Xiaoqing X Janda Kim D KD Cohen Seth M SM

Chemical communications (Cambridge, England) 20130312 31

Tropolone emerged from the screening of a chelator fragment library (CFL) as an inhibitor of the Zn(2+)-dependent virulence factor, Pseudomonas aeruginosa elastase (LasB). Based on this initial hit, a series of substituted tropolone-based LasB inhibitors was prepared, and a compound displaying potent activity in vitro and in a bacterial swarming assay was identified. Importantly, this inhibitor was found to be specific for LasB over other metalloenzymes, validating the usage of tropolone as a vi ...[more]

PMID: 23482955

Similar Datasets

Project description:Metal-catalyzed lipid oxidation is a major factor in food waste, as it reduces shelf life. Addressing this issue, our study investigates the potential of hydrolysates derived from potato protein, a by-product of potato starch production, as metal chelating antioxidants. Through sequential en-zymatic hydrolysis using Alcalase or Trypsin combined with Flavourzyme, we produced various hydrolysates, which were then fractionated via ultrafiltration. Using a combination of peptidomics and bioinformatics, we predicted the presence of metal-chelating and free radical scavenging pep-tides across all hydrolysate fractions, with a trend indicating a higher content of antioxidant pep-tides in lower molecular weight fractions. To validate these predictions, we utilized Surface Plas-mon Resonance (SPR) and a 9-day emulsion storage experiment. While SPR demonstrated poten-tial in identifying antioxidant activity, it faced challenges in differentiating between hydrolysate fractions due to significant standard errors. In the storage experiment, all hydrolysates showed li-pid oxidation inhibition, though not as effectively as EDTA. Remarkably, one fraction (AF13) was not significantly different (p < 0.05) from EDTA in suppressing hexanal formation. These results highlight SPR and peptidomics/bioinformatics as promising yet limited methods for antioxidant screening. Importantly, this study reveals the potential of potato protein hydrolysates as antioxi-dants in food products, warranting further research. The aim of this study was to evaluate the potential of SPR as a screening technique for potato protein hydrolysates (PPH) as metal chelating antioxidants. More specifically, the goal was to correlate the KD of the hydrolysates, determined by SPR, with development of different oxidation markers obtained during a storage experiment where the PPHs were added as antioxidants to 5 % fish oil-in-water emulsions at pH 7. Additionally, the effect of using different combinations of enzymes on antioxidant activity of the hydrolysates was investigated and peptidomics was applied to characterize the composition of the hydroly-sates. With the aim of achieving shorter peptides through a high degree of hydrolysis (DH%), this study utilized sequential hydrolysis, where Trypsin (Try) or Alcalase (Alc) was paired with Flavourzyme (Fla). Owing to its broad specificity as a heterogeneous combination of various endo- and exopeptidases [20], Flavourzyme is known to facilitate high DH% and the production of short peptides [21]. However, it has been reported that when used solely with a denatured substrate protein of limited solubility, Flavourzyme can yield a significantly lower DH% than anticipated [19]. Consequently, this study was designed to include substrate pre-digestion prior to Flavourzyme addition.

Project description:Mitochondrial Fe-S cluster biosynthesis is accomplished within yeast utilizing the biophysical attributes of the "Isu1" scaffold assembly protein. As a member of a highly homologous protein family, Isu1 has sequence conservation between orthologs and a conserved ability to assemble [2Fe-2S] clusters. Regardless of species, scaffold orthologs have been shown to exist in both "disordered" and "structured" conformations, a structural architecture that is directly related to conformations utilized during Fe-S cluster assembly. During assembly, the scaffold helps direct the delivery and utilization of Fe(ii) and persulfide substrates to produce [2Fe-2S] clusters, however Zn(ii) binding alters the activity of the scaffold while at the same time stabilizes the protein in its structured state. Additional studies confirm Zn binds to the scaffold's Cys rich active site, and has an impact on the protein's ability to make Fe-S clusters. Understanding the interplay between Fe(ii) and Zn(ii) binding to Isu1 in vitro may help clarify metal loading events that occur during Fe-S cluster assembly in vivo. Here we determine the metal?:?protein stoichiometry for Isu1 Zn and Fe binding to be 1?:?1 and 2?:?1, respectively. As expected, while Zn binding shifts the Isu1 to its structured state, folding is not influenced by Fe(ii) binding. X-ray absorption spectroscopy (XAS) confirms Zn(ii) binds to the scaffold's cysteine rich active site but Fe(ii) binds at a location distinct from the active site. XAS results show Isu1 binding initially of either Fe(ii) or Zn(ii) does not significantly perturb the metal site structure of alternate metal. XAS confirmed that four scaffold orthologs bind iron as high-spin Fe(ii) at a site composed of ca. 6 oxygen and nitrogen nearest neighbor ligands. Finally, in our report Zn binding dramatically reduces the Fe-S cluster assembly activity of Isu1 even in the presence of frataxin. Given the Fe-binding activity we report for Isu1 and its orthologs here, a possible mechanism involving Fe(ii) transport to the scaffold's active site during cluster assembly has been considered.

Dataset Information

Antagonism of a zinc metalloprotease using a unique metal-chelating scaffold: tropolones as inhibitors of P. aeruginosa elastase.

Publications

Antagonism of a zinc metalloprotease using a unique metal-chelating scaffold: tropolones as inhibitors of P. aeruginosa elastase.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets