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Cloning and expression in Escherichia coli of cDNA encoding house dust mite allergen Der f 3, serine protease from Dermatophagoides farinae
Author(s) -
Nishiyama Chiharu,
Yasuhara Takaomi,
Yuuki Toshifumi,
Okumura Yasushi
Publication year - 1995
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(95)01291-5
Subject(s) - complementary dna , microbiology and biotechnology , serine protease , biochemistry , escherichia coli , protease , peptide sequence , amino acid , biology , serine , house dust mite , chemistry , allergen , enzyme , gene , allergy , immunology
Der f 3 is one of the allergens produced by house dust mite Dermatophagoides farinae showing serine protease activity. Based on its amino acid sequence, a cDNA clone encoding Der f 3 was isolated from a cDNA library of D. farinae . Sequencing analysis of the clone revealed the presence of an open reading frame of 780 bp, which encodes a mature protein of 232 amino acids with 27 amino acids of pre‐pro sequence at the N‐terminus. When proDer f 3 was produced in Escherichia coli as a fused protein with glutathione‐ S ‐transferase, the fused protein was accumulated as inclusion bodies. The protein purified with 8 M urea and glutathione‐affinity column chromatography, however, did not show protease activity. When an arginine residue was introduced at the C‐terminus of the pro‐region in place of threonine, removal of the pro‐region to produce an active mature protease was observed. The specificity and the activity of this recombinant protease were almost the same as those of native Der f 3.