Cryoprotective effect of an insect antifreeze protein MpAFP 698 and its mutants from the desert beetle Microdera punctipennis

An insect antifreeze protein, MpAFP698, from the desert beetle Microdera punctipennis, shares 77% similarity with TmTHP (YL-3) from Tenebrio molitor. The predicted tertiary structure for MpAFP698 was modeled as a regular A-helix with TCT motifs arrayed to form the putative ice-binding surface. This model was validated via site-directed mutagenesis. The results suggest that the desert beetle in central Asia has evolved antifreeze proteins similar to those from North America continent. In an additional study, the recombinant MpAFP698 and its mutants were expressed in Escherichia coli strain BL21 (DE3). The thermal hysteresis activity of MpAFP698 was 1.73 degree C at 1.0 mg/ml. MpAFP698 was also tested by in vitro antifreeze activity assay to evaluate its cryoprotective effect at -20 degree C. MpAFP698 displays high cryoprotective effect on bacteria cells at freezing temperatures.