Ochratoxin binding to phenylalanyl-tRNA synthetase: computational approach to the mechanism of ochratoxicosis and its antagonism.

Ochratoxin A (OA) is a toxic isocoumarin derivative released by various species of mold which grow on grain, coffee, and nuts, representing a serious worldwide health problem. Among other mechanisms of toxicity, it has been suggested that OA inhibits phenylalanyl-tRNA synthetase (PheRS), thereby reducing protein synthesis. Using the crystal structure of PheRS from Thermusthermophilus, we have modeled its interactions with OA as well as with phenylalanyl adenylate (FAMP), the high-affinity intermediate substrate of PheRS. Our results indicate that while OA may be capable of weakly inhibiting PheRS, the OA-PheRS complex cannot adopt the same conformation as does FAMP-PheRS, contrary to previous assumptions. Relative to FAMP, the phenylalanyl moiety is found to bind more shallowly and in a different overall conformation. Free-energy perturbation calculations of the relative free energies of binding of OA with the phenolic moiety protonated versus deprotonated suggest that the protonated form binds significantly more strongly. Two alternative binding modes were also identified which cannot be discounted on the basis of these calculations. Our results, however, do not suggest binding stronger than millimolar for any of the binding modes, a conclusion which is in agreement with more recent experimental findings. This, in turn, suggests that the previously observed antagonistic effects of aspartame and piroxicam are more likely due to their prevention of OA binding to human serum albumin than to PheRS, which is in agreement with binding studies as well as with preliminary simulations performed in our laboratory.