title: Characterization of a secreted cystatin from the tick Rhipicephalus haemaphysaloides.
abstract: A novel cystatin, designated RHcyst-2, was isolated from the tick Rhipicephalus haemaphysaloides. The full-length cDNA of RHcyst-2 is 773 bp, including an intact open reading frame encoding an expected protein of 139 amino acids and consisting of a 23 amino acids signal peptide. Predicted RHcyst-2 mature protein molecular weight is about 13 kDa, isoelectric point is 4.96. A sequence analysis showed that it has significant homology with the known type 2 cystatins. The recombinant protein of RHcyst-2 was expressed in a glutathione S-transferase-fused soluble form in Escherichia coli, and its inhibitory activity against cathepsin L, B, C, H, and S, as well as papain, was identified by fluorogenic substrate analysis. The results showed that rRHcyst-2 can effectively inhibit the six cysteine proteases' enzyme activities. An investigation of the RHcyst-2 genes' expression profile by quantitative reverse transcription-PCR demonstrated that it was more richly transcribed in the embryo (egg) stage and mainly distributed in the mid-gut of adult ticks. Western blot analysis confirmed that RHcyst-2 was secreted into tick saliva.
authors.affiliation: Key Laboratory of Animal Parasitology of Ministry of Agriculture, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai, 200241, China.
authors.forename: Yujian
authors.initials: Y
authors.lastname: Wang
authors.forename: Xinmao
authors.initials: X
authors.lastname: Yu
authors.forename: Jie
authors.initials: J
authors.lastname: Cao
authors.forename: Yongzhi
authors.initials: Y
authors.lastname: Zhou
authors.forename: Haiyan
authors.initials: H
authors.lastname: Gong
authors.forename: Houshuang
authors.initials: H
authors.lastname: Zhang
authors.forename: Xiangrui
authors.initials: X
authors.lastname: Li
authors.forename: Jinlin
authors.initials: J
authors.lastname: Zhou
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D060829:Arthropod Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D015891:Cystatins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007814:Larva / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009758:Nymph / Q000235:genetics / Q000378:metabolism; D010063:Ovum / Q000378:metabolism; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D048494:Rhipicephalus / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D016415:Sequence Alignment; D014018:Tissue Distribution
chemical_list: D060829:Arthropod Proteins; D015891:Cystatins; D018076:DNA, Complementary; D012333:RNA, Messenger