Ochratoxin binding to phenylalanyl-tRNA synthetase: Computational approachto the mechanism of ochratoxicosis and its antagonism

Ochratoxin A (OA) is a toxic isocoumarin derivative released by various spe cies of mold which grow on grain, coffee, and nuts, representing a serious worldwide health problem. Among other mechanisms of toxicity, it has been s uggested that OA inhibits phenylalanyl-tRNA synthetase (PheRS), thereby red ucing protein synthesis. Using the crystal structure of PheRS from Thermus thermophilus, we have modeled its interactions with OA as well as with phen ylalanyl adenylate (FAMP), the high-affinity intermediate substrate of PheR S. Our results indicate that while OA may be capable of weakly inhibiting P heRS, 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 conforma tion. Free-energy perturbation calculations of the relative free energies o f binding of OA with the phenolic moiety protonated versus deprotonated sug gest that the protonated form binds significantly more strongly. Two altern ative binding modes were also identified which cannot be discounted on the basis of these calculations. Our results, however, do not suggest binding s tronger than millimolar for any of the binding modes, a conclusion which is in agreement with more recent experimental findings. This, in turn, sugges ts that the previously observed antagonistic effects of aspartame and pirox icam are more likely due to their prevention of OA binding to human serum a lbumin than to PheRS, which is in agreement with binding studies as well as with preliminary simulations performed in our laboratory.