Purification and Characterization of Aspartic Protease Derived from Sf9 Insect Cells

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概要

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• An aspartic protease that is significantly produced by baculovirus-infected Spodoptera frugiperda Sf9 insect cells was purified to homogeneity from a growth medium. To monitor aspartic protease activity, an internally quenched fluoresce (IQF) substrate specific to cathepsin D was used. The purified aspartic protease showed a single protein band on SDS–PAGE with an apparent molecular mass of 40 kDa. The N-terminal amino acid sequence of the enzyme had a high homology to a Bombyx mori aspartic protease. The enzyme showed greatest affinity for the IQF substrate at pH 3.0 with a Km of 0.85 μM. The kcat and kcat⁄Km values were 13 s−1 and 15 s−1 μM−1 respectively. Pepstatin A proved to be a potent competitive inhibitor with inhibitor constant, Ki, of 25 pM.

• 社団法人 日本農芸化学会の論文

著者

• Gotoh Takeshi

  Department of Engineering in Applied Chemistry, Akita University

• Kikuchi Ken-Ichi

  Department of Engineering in Applied Chemistry, Akita University

• TAKAHASHI Saori

  Akita Research Institute of Food and Brewing

• Ono Hiroki

  Department of Materials-Process Engineering and Applied Chemistry for Environments, Akita University

• NIRASAWA Satoru

  Japan International Research Center for Agricultural Sciences

• Gotoh Takeshi

  Department Of Engineering In Applied Chemistry Graduate School Of Engineering And Resource Science A

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