Purification and characterisation of blarinasin, a new tissue kallikrein-like protease from the short-tailed shrew Blarina brevicauda: comparative studies with blarina toxin

• Published in: Biological chemistry Vol. 386; no. 2; pp. 177 - 182
• Main Authors: Kita, Masaki, Okumura, Yuushi, Ohdachi, Satoshi D., Oba, Yuichi, Yoshikuni, Michiyasu, Nakamura, Yasuo, Kido, Hiroshi, Uemura, Daisuke
• Format: Journal Article
• Language: English
• Published: Germany Walter de Gruyter 01.02.2005
• Subjects: Amino Acid Sequence; amino acid sequence analysis; Animals; Base Sequence; Blarina; Blarina brevicauda; blarina toxin; blarinasin; Hydrogen-Ion Concentration; Kallikreins - chemistry; Kallikreins - isolation & purification; Molecular Sequence Data; Protease Inhibitors - pharmacology; salivary glands; Sequence Alignment; short-tailed shrew; Shrews; Substrate Specificity; tissue kallikrein; Tissue Kallikreins - chemistry
• ISSN: 1431-6730; 1437-4315
• DOI: 10.1515/BC.2005.022

A new tissue kallikrein-like protease, blarinasin, has been purified from the salivary glands of the short-tailed shrew Blarina brevicauda. Blarinasin is a 32-kDa N-glycosylated protease with isoelectric values ranging between 5.3 and 5.7, and an optimum pH of 8.5 for enzyme activity. The cloned blarinasin cDNA coded for a pre-pro-sequence and a mature peptide of 252 amino acids with a catalytic triad typical for serine proteases and 43.7–54.0% identity to other mammalian tissue kallikreins. Blarinasin preferentially hydrolysed Pro-Phe-Arg-4-methylcoumaryl-7-amide (MCA) and N-tert-butyloxycarbonyl-Val-Leu-Lys-MCA, and preferentially converted human high-molecular-weight kininogen (HK) to bradykinin. The activity of blarinasin was prominently inhibited by aprotinin (K i=3.4 nM). A similar kallikrein-like protease, the lethal venom blarina toxin, has previously been purified from the salivary glands of the shrew Blarina and shows 67.9% identity to blarinasin. However, blarinasin was not toxic in mice. Blarinasin is a very abundant kallikrein-like protease and represents 70–75% of kallikrein-like enzymes in the salivary gland of B. brevicauda.