Structure of the Ochratoxin A Binding Site within Human Serum Albumin

Abstract

The binding site of ochratoxin A (OTA) within domain 2A of human serum albumin (HSA) is examined by theoretical simulations and site-directed mutagenesis experiments. The calculated binding constant, based on docking experiments and theoretical affinity constants derived from the empirical free energy of binding as implemented in AutoDock 3.0, for the OTA dianion (3.7 × 10 6 M -1 ) is in good agreement with experimental value of 5.2 × 10 6 M -1 . The carboxy terminus of OTA associates with R218 and R222 of the protein. Binding is reduced by over an order of magnitude for the mutant R218A in both experiments and theoretical simulations. The carbonyl of the lactone and the phenolic group of OTA are in close proximity to R257. The experimental binding constant of OTA to the R257A mutant is 1.6 × 10 5 M -1 , over an order of magnitude smaller than for the wild-type protein. The predicted binding constant based on a comparison of the lowest-energy conformer from docking studies performed in AutoDock 3.0 of OTA to the R257A mutant (8.3 × 10 4 M -1 ) is also in good agreement with the experimental result. R257 clearly plays an important role in the binding of the isocoumarin ring of OTA by serving as a proton acceptor and stabilizing the binding through the creation of an ion pair with the phenoxide group on OTA.

Binding of Warfarin Influences the Acid‐Base Equilibrium of H242 in Sudlow Site I of Human Serum Albumin

“…The cocrystallized structure of W was removed prior to AutoDock 3.0 analysis. Details of the parameters used in the calculation are described elsewhere (10).…”

“…In a recent series of articles, the details of the binding of the fungal mycotoxin ochratoxin A (OTA; Chart 1) to Site I of HSA was examined by optical spectroscopy, isothermal titration calorimetry (ITC), genetic modification of HSA and molecular modeling (6–10). One of the most interesting details associated with the OTA‐HSA complex is the effect of binding on the p K a of the hydroxyl group.…”

Flavonoid aglycones can compete with Ochratoxin A for human serum albumin: A new possible mode of action

“…On human serum albumin (HSA) the primary binding site is located in subdomain IIA, in site I [11,12]. The bioavailability is about 93%, the half-life of OTA in plasma is around 1 month in humans and more than 99.8% of the circulating toxin is bound to albumin [13,14].…”

Ochratoxin A in human blood serum – retrospective long-term data

“…The reference range for human serum albumin (HSA) concentrations in blood is 30-50 mg/ml, and it has a relative molecular mass of 67,000. Consequently, OTA concentrations theoretically could reach levels of up to 360-600 µg OTA per ml of blood, enabling passive absorption from the digestive system even when OTA concentration in plasma is higher than that in the intestine (Kumagai 1988;Il'ichev et al 2002;Perry et al 2004). Another aspect related to HSA binding of OTA is its long persistence in blood serum, making it uniquely useful as a biomarker of exposure (Scott 2005).…”