Modification of the Functional Arginine Residue in Soybean Trypsin Inhibitor (Kunitz) by Immobilized Peptidylarginine Deiminase(Biological Chemistry)
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概要
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We immobilized peptidylarginine deiminase, a Ca^<2+>-dependent protein-modifying enzyme which catalyzes the deimination of arginyl residues in protein, on formyl-Sepharose 4B by reductive amination. the immobilized enzyme was quite stable in the presence of reducing reagents and chelators such as DTT and EGTA. The immobilized enzyme was inactivated under the catalytic conditions in the absence of substrate. the inactive enzyme, however, was totally reactivated by replacement of Ca^<2+> by am mixture of reducing reagent and chelator in the equilibration buffer. Our preceding paper described the specific modification of the functional arginine residue in soybean trypsin inhibitor (Kunitz) (STI) by peptidylarginine deiminase. In this work, the rate of the modification of STI by the immobilized enzyme packed in a column was measured by continuous application of STI at constant temperature and flow rate. the extent of the modification of STI put on the column was linear to about 70% with the reciprocal of the flow rate. The effects of pH and calcium ion on the catalytic behavior of the immobilized enzyme were essentially the same as those of the soluble enzyme reported previously. the steady-state kinetic analyses of STI modification by the immobilized enzyme found that the Km and к_<cat>/Km values were 6.0〜8.8 μM and 8.7×10^4M^<-1>・S^<-1>(37℃), respectively and the activation energy was 10.2〜15.7 kcal/mol.
- 社団法人日本農芸化学会の論文
- 1987-02-23
著者
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Sugawara Kiyoshi
Laboratories of Biochemistry, lbaraki University
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Takahara Hidenari
Laboratories of Biochemistry, lbaraki University
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Asahi Nobuo
Laboratory Of Biochemistry Department Of Agricultural Chemistry Ibaraki University
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- Isolation and Molecular Cloning of Epidermal- and Hair Follicle-Specific Peptidylarginine Deiminase (Type III) from Rat^1
- Modification of the Functional Arginine Residue in Soybean Trypsin Inhibitor (Kunitz) by Immobilized Peptidylarginine Deiminase(Biological Chemistry)
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