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j.enzmictec.2011.04.019.pdf | 362.54 kB | Adobe PDF | 見る/開く |
タイトル: | Effects of amines and aminoalcohols on bovine intestine alkaline phosphatase activity |
著者: | Sekiguchi, Satoshi Hashida, Yasuhiko Yasukawa, Kiyoshi Inouye, Kuniyo |
著者名の別形: | 井上, 國世 |
キーワード: | Alkaline phosphatase Amine Aminoalcohol Bovine intestine alkaline phosphatase Enzyme immunoassay |
発行日: | Jul-2011 |
出版者: | Elsevier Inc. |
誌名: | Enzyme and Microbial Technology |
巻: | 49 |
号: | 2 |
開始ページ: | 171 |
終了ページ: | 176 |
抄録: | Bovine intestine alkaline phosphatase (BIALP) is widely used as a signaling enzyme in sensitive assays such as enzyme immunoassay (EIA). In this study, we evaluated the effects of various aminoalcohols and amines on the activity of BIALP in the hydrolysis of p-nitrophenyl phosphate (pNPP) at pH 9.8, at 20 °C. The Kcat values at 0.05 M diethanolamine, 0.1 M triethanolamine, and 0.2 M N-methylethanolamine were 190 ± 10, 840 ± 30, and 500 ± 10 s[−1], respectively. The Kcat values increased with increasing concentrations of diethanolamine, triethanolamine, and N-methylethanolamine and reached 1240 ± 60, 1450 ± 30, and 2250 ± 80 s[−1], respectively, at 1.0 M. On the other hand, the Kcat values at 0.05-1.0 M ethanolamine, ethylamine, methylamine, and dimethylamine were in the range of 100–600 s[−1]. These results indicate that diethanolamine, triethanolamine and N-methylethanolamine highly activate BIALP and might be suitable as a dilution buffer of BIALP in EIA. Interestingly, the Km values increased with increasing concentrations of diethanolamine and N-methylethanolamine, but not triethanolamine: the Km value at 1.0 M diethanolamine (0.83 ± 0.15 mM) was 12-fold higher than that at 0.05 M (0.07 ± 0.01 mM), and that at 1.0 M N-methylethanolamine (2.53 ± 0.20 mM) was 14-fold higher than that at 0.2 M (0.18 ± 0.02 mM), while that at 1.0 M triethanolamine (0.31 ± 0.01 mM) was similar as that at 0.2 M (0.25 ± 0.01 mM), suggesting that the mechanisms of BIALP activation are different between the aminoalcohols. |
著作権等: | © 2011 Elsevier Inc. This is not the published version. Please cite only the published version. この論文は出版社版でありません。引用の際には出版社版をご確認ご利用ください。 |
URI: | http://hdl.handle.net/2433/143675 |
DOI(出版社版): | 10.1016/j.enzmictec.2011.04.019 |
PubMed ID: | 22112405 |
出現コレクション: | 学術雑誌掲載論文等 |
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