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KMID : 0359320030430010145
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Korean Journal of Veterinary Research
2003 Volume.43 No. 1 p.145 ~ p.150
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A simple and sensitive assay for chitinolytic activity of the recombinant CHT1 proteins from the hard tick H. longicornis using ethylene glycol chitin
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You Myung-Jo
Kozo Fujisaki
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Abstract
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To determine effectively the chitinolytic activity of rCHT1 from the hard tick H. longicornis expressed in baculovirus-mediated Spodoptera frugiperda (Sf) 9 cells, a simple and sensitive assay system was established in slid phase using agarose gel containing ethylene glycol chitin as substrate. The various factors affecting the efficacy of the assay were also investigated. The effects of various temperature, dosages of proteins, pH of media and time course of reaction were examined to verify the sensitivity of assay for chitinolytic activity of rCHT1 protein. It was found that the optimal reactive conditions were 37˚C of temperature, 12 to 15 hours of reactive times, 0.1 ㎍ of protein concentration and pH 5 to 7 of media. Using the assay system designed, the functional activities of H. longicornis rCHT1 protein could be evaluated simply and sensitively.
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