Binding nature and denaturation of protein during interaction with galloylglucose.

Analysis of insoluble complexes between tetragalloylglucose and proteins following a series of successive washes with buffer indicated (1) heterogeneity of binding between galloylglucose and protein and (2) irreversible denaturation of protein during interaction with galloylglucose. Relatively large amounts of tetragalloylglucose were removed by initial washes, indicating weak, low affinity binding, whereas smaller amounts removed by subsequent washes suggest bonds with a higher affinity. Although the maximum number of bindings sites, calculated per 10,000 M(r) of protein, was similar for BSA, myoglobin and lysozyme, the proportion of these sites that appeared to have high affinity, varied from 8 to 29%. The low proportion of strongly binding sites in lysozyme explains its relatively low tannin-complexing ability. Solubility decrease in protein during successive washing and decrease in the beta-glucosidase activity indicate that irreversible denaturation of protein occurs, which progresses with an increase in the incubation time with galloylglucose and galloylglucose/protein molar ratio in the mixture. Relative affinity of galloylglucose is directly related to the ability to cause irreversible denaturation.