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Production of high concentrations of fuel ethanol by fermentation of ligneous biomass

木质生物质发酵生产高浓度燃料乙醇

基本信息

批准号：
18580332
负责人：
OHTA Kazuyoshi
金额：
$ 2.41万
依托单位：
University of Miyazaki
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18580332/
关键词：
Penicillium citrinum Aspergillus japonicus xylan xylanase β-xylosidase filamentous fungi biomass ethanol xylan xylanase β-xylosidase filamentous fungi biomass ethanol 発酵大腸菌

项目摘要

1)An extracellular endo-1,4-β-xylanase was purified from the culture filtrate of a filamentous fungus, Penicillium citrinum strain FERM P-15944. The purified enzyme showed a single band on SDS-PAGE with an apparent M_r of 31.6 kDa. An open reading frame of the xylanase gene ^<xynB was interrupted by nine introns and encoded a presumed prepropeptide of 25 amino acids and a mature protein of 302 amino acids. Sequence alignment showed that the P. citrinum enzyme belongs to glycoside hydrolase family 10.2)An extracellular protein exhibiting β-xylosidase activity was purified from the culture filtrate of a filamentous fungus Aspergillus japonicus strain MU-2. The purified enzyme was a glycoprotein with an apparent M_r of 113.2 kDa The enzyme also had hydrolytic activities toward p-nitropheny1-β-D-glucopyranoside and p-nitropheny1-β-L-arabinofuranoside. An open reading frame of the β-xylo, sidase gene ^<xyl>A, consisting of 2412 bp, was not interrupted by introns, and it encoded a presumed s … More ignal peptide of 17 amino acids and a mature protein of 787 amino acids. The deduced amino acid sequence showed a high degree of identity (69%)to Aspergillus nigerβ-xylosidase XlnD that belongs to glycoside hydrolase family 3.3)An extracellular xylanase from A. japonicus was purified with a yield of 28.7% of the activity and a 21-fold increase in specific activity. The xylanase showed a single band on SDS-PAGE with an apparent M_r of 25.1 kDa. A pair of degenerate PCR primers was synthesized according to the internal amino acid sequence (EDYGEYN)and a highly conserved sequence (NHFNAWA)among GH family-11 xylanases from Aspergillus spp. Two distinct internal sequences were amplified from genomic DNA of A. japonicus as a template by PCR with the primer pair. Two genomic DNA segments encoding xylanase were cloned using two DIG-labeled amplified fragments The deduced amino acid sequences of two xylanase genes, ^<xynG1 and ^<xynG2, revealed that purified xylanase corresponded to ^<xynG2 gene product. Less

1)从丝状真菌柑橘青霉菌株FERM P-15944的培养滤液中纯化胞外内切1,4-β-木聚糖酶。纯化的酶在 SDS-PAGE 上显示单条带，表观 M_r 为 31.6 kDa。木聚糖酶基因^<xynB的开放阅读框被9个内含子中断，并编码25个氨基酸的假定前原肽和302个氨基酸的成熟蛋白质。序列比对显示P. citrinum酶属于糖苷水解酶家族10.2)从丝状真菌日本曲霉菌株MU-2的培养滤液中纯化出具有β-木糖苷酶活性的胞外蛋白。纯化的酶是一种糖蛋白，表观M_r为113.2 kDa。该酶还具有对硝基苯基1-β-D-吡喃葡萄糖苷和对硝基苯基1-β-L-阿拉伯呋喃糖苷的水解活性。 β-木糖苷酶基因 ^<xyl>A 的开放阅读框由 2412 bp 组成，未被内含子打断，它编码了一个由 17 个氨基酸组成的假定信号肽和一个由 787 个氨基酸组成的成熟蛋白。推导的氨基酸序列与属于糖苷水解酶家族的黑曲霉β-木糖苷酶XlnD具有高度的一致性(69%)。3.3)纯化了日本曲霉的胞外木聚糖酶，其活性产率为28.7%，比活性增加了21倍。木聚糖酶在 SDS-PAGE 上显示单条带，表观 M_r 为 25.1 kDa。根据曲霉属GH家族11木聚糖酶内部氨基酸序列(EDYGEYN)和高度保守序列(NHFNAWA)合成一对简并PCR引物。以刺参基因组DNA为模板，用引物对通过PCR扩增出两条不同的内部序列。使用两个DIG标记的扩增片段克隆编码木聚糖酶的两个基因组DNA片段。两个木聚糖酶基因的推导的氨基酸序列，^<xynG1和^<xynG2，揭示纯化的木聚糖酶对应于^<xynG2基因产物。较少的