Enhancement effect of solutes of low molecular mass on the insect antifreeze protein ApAFP752 from Anatolica polita

Many ectotherms produce antifreeze proteins (AFPs), also known as thermal hysteresis proteins that can lower non-colligatively freezing point of body liquids without altering the melting point. The difference between the depressed freezing point and the melting point, termed thermal hysteresis (TH), is usually a measure of the antifreeze activity of AFPs. Certain low molecular mass molecules and proteins can further enhance the antifreeze activity of AFPs. The enhancement effects of several salts, polycarboxylates, and polyhydroxy compounds on an antifreeze protein (ApAFP752) from the desert beetle Anatolica polita were determined using differential scanning calorimetry. These observations suggest that the salts including NaCl, KCl, CaCl2, and MgCl2 can enhance colligatively the thermal hysteresis activity (THA) of ApAFP752. The bivalent ions were more effective than monovalent ions at low concentrations. The enhancing effects of three polycarboxylates were higher than that of the neutral salts, and the more carboxylate groups per enhancer molecule the better the efficiency of the enhancer. Ethylenediaminetetraacetic acid disodium salt showed the strongest synergism ability and increased the THA nearly 13-fold from 0.45 °C in its absence to 5.67 °C. Glycerol and trehalose, the polyhydroxy compounds studied, had limited enhancement efficiency on ApAFP752, and only enhance the TH of the protein at high concentration. Mechanisms are discussed.