Alessio Amadasi

Design of O-acetylserine sulfhydrylase inhibitors by mimicking Nature

The inhibition of cysteine biosynthesis in prokaryotes and protozoa has been proposed to be relevant for the development of antibiotics. Haemophilus influenzae O-acetylserine sulfhydrylase (OASS), catalyzing l-cysteine formation, is inhibited by the insertion of the C-terminal pentapeptide (MNLNI) of serine acetyltransferase into the active site. Four-hundred MNXXI pentapeptides were generated in silico, docked into OASS active site using GOLD, and scored with HINT. The terminal P5 Ile accounts for about 50% of the binding energy. Glu or Asp at position P4 and, to a lesser extent, at position P3 also significantly contribute to the binding interaction. The predicted affinity of 14 selected pentapeptides correlated well with the experimentally determined dissociation constants. The X-ray structure of three high affinity pentapeptide-OASS complexes were compared with the docked poses. These results, combined with a GRID analysis of the active site, allowed us to define a pharmacophoric scaffold for the design of peptidomimetic inhibitors.

Identification of Xenoestrogens in Food Additives by an Integrated in Silico and in Vitro Approach

In the search for xenoestrogens within food additives, we have analyzed the Joint FAO-WHO expert committee database, containing 1500 compounds, using an integrated in silico and in vitro approach. This analysis identified 31 potential estrogen receptor alpha ligands that were reduced to 13 upon applying a stringent filter based on ligand volume and binding mode. Among the 13 potential xenoestrogens, four were already known to exhibit an estrogenic activity, and the other nine were assayed in vitro, determining the binding affinity to the receptor and biological effects. Propyl gallate was found to act as an antagonist, and 4-hexylresorcinol was found to act as a potent transactivator; both ligands were active at nanomolar concentrations, as predicted by the in silico analysis. Some caution should be issued for the use of propyl gallate and 4-hexylresorcinol as food additives.

Pyridoxal 5′-Phosphate-Dependent Enzymes: Catalysis, Conformation, and Genomics

Pyridoxal 5′-phosphate (PLP) is the biologically active form of B6 vitamers. PLP plays the role as coenzyme in more than 160 different enzymes acting on a variety of substrates and inhibitors predominantly possessing amine and carboxylate moieties. The key feature of PLP-dependent enzymes is the catalytic versatility, carrying out transamination, decarboxylation, and beta and gamma elimination and substitution via several catalytic intermediates. Catalysis frequently accompanies a conformational transition between an open, less active state and a closed, fully active state. Recent investigations on functional genomics of PLP-dependent enzymes and the search of novel targets for therapeutic activity indicate that this class of enzymes has not been fully explored in the past and calls for more intensive and focussed studies.