Studies on the Purification and Properties of N-acetyl-β-D-glucosaminidase from Cabbage
碩士 === 靜宜大學 === 食品營養學系 === 80 === N-acetyl-β-D-glucosaminidase was purified from cabbage by sequential steps of buffer extraction,ammonium sulfate fractionation. Sephadex G-75 gel filtration, chromatofocusing, and Sepharose CL-6B gel filtration. By these steps, the purity of the enzyme increased by...
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ndltd-TW-080PU0002550032019-05-15T23:17:14Z http://ndltd.ncl.edu.tw/handle/wuxq39 Studies on the Purification and Properties of N-acetyl-β-D-glucosaminidase from Cabbage 甘藍β-N-乙醯胺基葡萄糖苷酶之純化與性質研究 Young, Fong-Ping 楊鳳平 碩士 靜宜大學 食品營養學系 80 N-acetyl-β-D-glucosaminidase was purified from cabbage by sequential steps of buffer extraction,ammonium sulfate fractionation. Sephadex G-75 gel filtration, chromatofocusing, and Sepharose CL-6B gel filtration. By these steps, the purity of the enzyme increased by 358 folds, the recovery of activity was 17.4%. The purified enzyme was homogeneous as examined by polyacrylamide gel electrophoresls. The purified N-acetyl-β-D-glucosaminidase had an optium pH of 4.0, optium temperature of 60°C, Km value of 0.94 mM, activation energy of 10.2 Kcal/mole, activation energy of thermal inactivation of 8.9 Kcal/mole for hydrolysis of p-nitrophenyl-N-acetyl-β-D-glucosaminde. The enzyme showed molecular weight of 151,000 as determined by Sepharose CL-6B gel filtration. One subunit with molecular weight of 21,000 was observed as determined by SDSpolyacrylamide gel electrophoresis. Heavy metalions(O.06mM) , Ag+ and Hg+2, significantly inhibited the activity of the enzyme. N-Bromosuccinimide (NBSI; O.06mM), phenylmethylsulfonyl fluoride (PMSF;O.6mM), and iodine (O.6mM) also inhibited the activity of the enzyme. The enzyme showed high activity for hydrolysis of p-nitrophenyl-N-acetyl-β-D-glucosaminide but had no action on p-nitrophenyl-N-acetyl-α-D-glucosaminide. The enzyme also hydrolyzed p-nitrophenyl-N-acetyl-β-D-galactosaminide and oligochitin (2-4)with rather low activity. From the chemical modification and Dixon-Webb kinetic studies, the amino acid residues, histidine, tryptophan , tyrosine, and serine are probably located at or near the activite site of the enzyme. Chang,Chen-Tien 張珍田 1992 學位論文 ; thesis 77 zh-TW |
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碩士 === 靜宜大學 === 食品營養學系 === 80 === N-acetyl-β-D-glucosaminidase was purified from cabbage by sequential steps of buffer extraction,ammonium sulfate fractionation. Sephadex G-75 gel filtration, chromatofocusing, and Sepharose CL-6B gel filtration. By these steps, the purity of the enzyme increased by 358 folds, the recovery of activity was 17.4%. The purified enzyme was homogeneous as examined by polyacrylamide gel electrophoresls.
The purified N-acetyl-β-D-glucosaminidase had an optium pH of 4.0, optium temperature of 60°C, Km value of 0.94 mM, activation energy of 10.2 Kcal/mole, activation energy of thermal inactivation of 8.9 Kcal/mole for hydrolysis of p-nitrophenyl-N-acetyl-β-D-glucosaminde. The enzyme showed molecular weight of 151,000 as determined by Sepharose CL-6B gel filtration. One subunit with molecular weight of 21,000 was observed as determined by SDSpolyacrylamide
gel electrophoresis. Heavy metalions(O.06mM) , Ag+ and Hg+2, significantly inhibited the activity of the enzyme. N-Bromosuccinimide (NBSI; O.06mM), phenylmethylsulfonyl fluoride (PMSF;O.6mM), and iodine (O.6mM) also inhibited the activity of the enzyme.
The enzyme showed high activity for hydrolysis of p-nitrophenyl-N-acetyl-β-D-glucosaminide but had no action on p-nitrophenyl-N-acetyl-α-D-glucosaminide.
The enzyme also hydrolyzed p-nitrophenyl-N-acetyl-β-D-galactosaminide and oligochitin (2-4)with rather low activity. From the chemical modification
and Dixon-Webb kinetic studies, the amino acid residues, histidine, tryptophan , tyrosine, and serine are probably located at or near the activite site of the enzyme.
|
author2 |
Chang,Chen-Tien |
author_facet |
Chang,Chen-Tien Young, Fong-Ping 楊鳳平 |
author |
Young, Fong-Ping 楊鳳平 |
spellingShingle |
Young, Fong-Ping 楊鳳平 Studies on the Purification and Properties of N-acetyl-β-D-glucosaminidase from Cabbage |
author_sort |
Young, Fong-Ping |
title |
Studies on the Purification and Properties of N-acetyl-β-D-glucosaminidase from Cabbage |
title_short |
Studies on the Purification and Properties of N-acetyl-β-D-glucosaminidase from Cabbage |
title_full |
Studies on the Purification and Properties of N-acetyl-β-D-glucosaminidase from Cabbage |
title_fullStr |
Studies on the Purification and Properties of N-acetyl-β-D-glucosaminidase from Cabbage |
title_full_unstemmed |
Studies on the Purification and Properties of N-acetyl-β-D-glucosaminidase from Cabbage |
title_sort |
studies on the purification and properties of n-acetyl-β-d-glucosaminidase from cabbage |
publishDate |
1992 |
url |
http://ndltd.ncl.edu.tw/handle/wuxq39 |
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