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Production of Useful Oligosaccharides by Lytic Enzymes

通过裂解酶生产有用的低聚糖

基本信息

批准号：
61560138
负责人：
USUI Taichi
金额：
$ 0.96万
依托单位：
Department of Agricultural Chemistry. Faculty of Agriculture Shizuoka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1986
资助国家：
日本
起止时间：
1986 至 1987
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-61560138/
关键词：
Lytic enzyme Lysozyme Chitinase Useful oligosaccharide by enzymatic synthesis 糖転移反応

项目摘要

Chitinase from the culture filtrates of Nocardia orientalis IFO 12806 or Trichoderma reesei was purified to apparent homogeneity by successive column chromatographies. The enzyme, which is essentially a hydrolase, also catalyzed a transglycosylation reaction on tetra-N-acetyl chitotetraose(GlcNAc)_4 and penta N-acetyl-chitopentaose(GLcNAc)_5. The enzyme converted the tetrasaccharide into hexa-N-acetyl-chitohexaose(GlcNAc)_6 and di-N-acetyl-chitobiose(GLcNAc)_2 as the major products. The corresponding values for penta-N-acetyl-chitopentaose(GLcNAc)_5 were hepta-N-acetyl-chitoheptaose(GLcNAc)_3. The rate of the transglycosylation depended on the temperature, the concentration of substrate and the pH.Transchitooligosylation from (GlcNAc)_5 to the 4-position of p-nitrophenyl 2-acetamido-2-deoxy-<beta>-D-glucopyranoside(PNP-GlcNAc) was efficiently induced through lysozyme catalysis in an aqueous solution containing methanol with a high concentration. Use of the aqueous methanol system in this reaction not only guaranteed solubility of PNP-GlcNAc substrate, but also resulted in a remarkable increase in p-nitrophenyl-penta-N-acetyl-<beta>-chitopentaoside(PNP-(GlcNAc)_5) production. PNP-(GlcNAc)_5 was substrate for lysozyme assay compared with p-nitrophenyl-tetra-N-acetyl-<beta>-chitotetraoside (PNP-(GlcNAc)_4).

从东方诺卡氏菌（Nocardia orientalis）IFO 12806或里氏木霉（Trichoderma reesei）的培养液中提取几丁质酶，经柱层析纯化至表观均一。该酶是一种水解酶，它也催化四-N-乙酰基壳四糖（GlcNAc）_4和五-N-乙酰基壳五糖（GlcNAc）_5的转糖基化反应。该酶将四糖转化为主要产物六-N-乙酰壳六糖（GlcNAc）_6和二-N-乙酰壳二糖（GlcNAc）_2。五-N-乙酰基-壳五糖（GLcNAc）_5的相应值为七-N-乙酰基-壳七糖（GLcNAc）_3。在<beta>高浓度甲醇水溶液中，利用溶菌酶的催化作用，可有效地将对硝基苯基2-乙酰氨基-2-脱氧-β-D-吡喃葡萄糖苷（PNP-GlcNAc）5转糖基至4位。在此反应中使用甲醇水溶液体系不仅保证了PNP-GlcNAc底物的溶解性，而且导致了对硝基苯基-五-N-乙酰基-β<beta>-壳戊糖苷（PNP-（GlcNAc）_5）产量的显著增加。PNP-（GlcNAc）_5与对硝基苯基-四-N-乙酰基-β<beta>-壳聚糖四糖苷（PNP-（GlcNAc）_4）相比，是溶菌酶活性测定的底物。