Trypsin-like protease of mites: purification and characterization of trypsin-like protease from mite faecal extract Dermatophagoides farinae. Relationship between trypsin-like protease and Der f III

Summary A serine protease from mite faecal extract, Dermatophagoides farinae, was purified using DEAE-Sephacel anion exchange chromatography and Supcrdex 75 pg gel chromato-graphy. The molecular weight of this protease was 34 kD on SDS-PAGE under reducing conditions. The optimal pH and temperature of the protease were 8-0 and 47 C, respectively. In addition, this protease cleaved arginyl or lysyl residue containing substrates selectively and was only inhibited by aprotinin, PUT-175, tind soy bean trypsin inhibitor and not by chymostatin, E-64 and iodoacetic acid. These results show that our purified serine protease belongs to the trypsin-type. Purified trypsin-like protease was shown to be allergenic by enzyme-linked immunosorbent assay. Antigeni-city of trypsin-like protease was completely different from those of Der f I and Der f II. Both, 20 N-terminal amino acid sequence and amino acid compositions of the purified protease were very similar to those of Der f III. Good similarities were found between trypsin-like protease and Der f III concerning physicochemical properties such as molecular weight on SDS-PAGE and ammonium sulphate solubility. Summarizing the above data, it can be concluded that a trypsin-like protease from mite faecal extract is actually the Der f III allergen and that it may be involved in the digestive process of the mite as it was found not in mite body but in mite faeces.