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CHARACTERIZATION OF A NEW KUNITZ‐TYPE SERINE PROTEASE INHIBITOR FROM THE HARD TICK Rhipicephalus hemaphysaloides
Author(s) -
Cao Jie,
Shi Lei,
Zhou Yongzhi,
Gao Xiao,
Zhang Houshuang,
Gong Haiyan,
Zhou Jinlin
Publication year - 2013
Publication title -
archives of insect biochemistry and physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.576
H-Index - 66
eISSN - 1520-6327
pISSN - 0739-4462
DOI - 10.1002/arch.21118
Subject(s) - serine protease , biology , microbiology and biotechnology , recombinant dna , kunitz sti protease inhibitor , protease inhibitor (pharmacology) , sf9 , trypsin inhibitor , protease , biochemistry , complementary dna , trypsin , spodoptera , enzyme , virology , virus , gene , antiretroviral therapy , viral load
A new K unitz‐type serine protease inhibitor, R hipilin‐2, was identified in the tick R hipicephalus hemaphysaloides. The c DNA sequence of R hipilin‐2 is 693 bp, and it encodes a deduced 195 amino acid protein with a size of 22 k D a. Bioinformatic analysis shows that R hipilin‐2 belongs to the K unitz‐type family of inhibitors, containing one K unitz domain with homology to the tissue factor pathway inhibitor. Using Real time polymerase chain reaction (Real time‐ PCR ), R hipilin‐2 m RNA transcripts were detected in tick salivary glands and midgut. Blood feeding induced transcript expression. The recombinant protein was expressed in insect S f9 cells and confirmed by immunofluorescence test and W estern blot analysis with an anti‐ H is antibody. The purified recombinant R hipilin‐2 inhibited serine protease trypsin and elastase, but not thrombin. The anticoagulant activity of R hipilin‐2 was shown by delaying normal clotting of rabbit plasma in the activated partial thromboplastin time tests. These results indicate that R hipilin‐2 is a novel K unitz‐type serine protease inhibitor involved in tick blood feeding.