標題: Mechanistic study of beta-xylosidase from Trichoderma koningii G-39
作者: Li, YK
Yao, HJ
Pan, IH
應用化學系
Department of Applied Chemistry
關鍵字: Bronsted plot;mechanism;secondary deuterium isotope effect;beta-xylosidase
公開日期: 1-二月-2000
摘要: The catalytic mechanism of the beta-xylosidase purified from the culture filtrate of Trichoderma Koningii G-39 was investigated. By NMR spectroscopy, the stereochemistry of the enzyme catalyzing the hydrolysis of 2,4,dinitrophenyl and p-nitrophenyl-beta-D-xylosides was found unequivocally to involve retention of the anomeric configuration, Based on the k(cat) values of a series of arylxylosides with leaving group pK(a)s in the range of 4-10, an extended Bronsted plot was constructed with a slope (beta(Ig)) near zero. Enzymatic hydrolysis of aryl-beta-D-xylosides in acetate buffer (pH 4.0) containing 3 or 5% methanol showed a constant product ratio (methylxyloside/xylose), indicating the presence of a common intermediate, probably the xylosyl-enzyme intermediate. In the presence of DTT, the k(cat) values of p-cyanophenyl-beta-D-xylopyranoside and p-nitrophenyl-beta-xylopyranoside increased greatly. A two-step mechanism involving the formation and breakdown of the xylosyl-enzyme intermediate was therefore proposed. The rate-limiting step is the breakdown of the intermediate. The secondary deuterium kinetic isotope effect (k(H)/k(D)) measured for 2,4-dinitrophenyl-beta-D-xyloside was 1.02+/-0.01, suggesting that the transition state for breakdown of the xylosyl-enzyme intermediate is S(N)2-like.
URI: http://hdl.handle.net/11536/30772
ISSN: 0021-924X
期刊: JOURNAL OF BIOCHEMISTRY
Volume: 127
Issue: 2
起始頁: 315
結束頁: 320
顯示於類別:期刊論文


文件中的檔案:

  1. 000085498500020.pdf

若為 zip 檔案,請下載檔案解壓縮後,用瀏覽器開啟資料夾中的 index.html 瀏覽全文。