Thermal stabilization of antithrombin III by sugars and sugar derivatives and the effects of nonenzymatic glycosylation.
A variety of neutral and acidic sugars and related compounds were evaluated in terms of their effect on the midpoint, Td, of the thermal denaturation curve of antithrombin III. The objectives were to determine which structural features of these molecules are responsible for their stabilizing properties and to identify more efficient stabilizers which combine the effects of lyotropic anions such as citrate with those of the polyols in a single molecule. The presence of one or more carboxylate groups in a sugar molecule invariably increased its stabilizing potency, whereas the number and position of hydroxyl groups appeared to have no influence on the molecules' stabilizing ability. Several compounds were shown to be effective in preserving antithrombin III activity during pasteurization for 10 h at 60 degrees C. However, the presence of reducing sugars invariably resulted in a decrease in activity following pasteurization, in spite of their ability to increase Td. In fact, when antithrombin III was pasteurized in the presence of 2 M glucose and 0.5 M citrate, it steadily lost its ability to inhibit thrombin even though Td under these conditions was 10 degrees C higher than in citrate alone where activity was preserved. This effect was shown to be coincident with the covalent incorporation of glucose into the protein molecule.
D000817:Anilino Naphthalenesulfonates; D000990:Antithrombin III / Q000378:metabolism*; D050260:Carbohydrate Metabolism; D003837:Deoxy Sugars / Q000378:metabolism; D003998:Dicarboxylic Acids / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D006358:Hot Temperature; D006801:Humans; D066298:In Vitro Techniques; D011489:Protein Denaturation; D012266:Ribose / Q000378:metabolism; D013237:Stereoisomerism; D013997:Time Factors
D000817:Anilino Naphthalenesulfonates; D003837:Deoxy Sugars; D003998:Dicarboxylic Acids; C027132:1-anilino-8-naphthalenesulfonate; D012266:Ribose; D000990:Antithrombin III; D005947:Glucose