CN108103047A - 一种高比活多功能纤维素酶突变体Nccle-mut及其编码基因和应用 - Google Patents

一种高比活多功能纤维素酶突变体Nccle-mut及其编码基因和应用 Download PDF

Info

Publication number
CN108103047A
CN108103047A CN201810085865.0A CN201810085865A CN108103047A CN 108103047 A CN108103047 A CN 108103047A CN 201810085865 A CN201810085865 A CN 201810085865A CN 108103047 A CN108103047 A CN 108103047A
Authority
CN
China
Prior art keywords
nccle
mut
mutant
gly
living
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
CN201810085865.0A
Other languages
English (en)
Inventor
刘丹妮
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Individual
Original Assignee
Individual
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Individual filed Critical Individual
Priority to CN201810085865.0A priority Critical patent/CN108103047A/zh
Publication of CN108103047A publication Critical patent/CN108103047A/zh
Pending legal-status Critical Current

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
    • C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
    • C12N9/2405—Glucanases
    • C12N9/2434—Glucanases acting on beta-1,4-glucosidic bonds
    • C12N9/2437—Cellulases (3.2.1.4; 3.2.1.74; 3.2.1.91; 3.2.1.150)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/70—Vectors or expression systems specially adapted for E. coli
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/80—Vectors or expression systems specially adapted for eukaryotic hosts for fungi
    • C12N15/81—Vectors or expression systems specially adapted for eukaryotic hosts for fungi for yeasts
    • C12N15/815—Vectors or expression systems specially adapted for eukaryotic hosts for fungi for yeasts for yeasts other than Saccharomyces
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y302/00—Hydrolases acting on glycosyl compounds, i.e. glycosylases (3.2)
    • C12Y302/01—Glycosidases, i.e. enzymes hydrolysing O- and S-glycosyl compounds (3.2.1)
    • C12Y302/01004—Cellulase (3.2.1.4), i.e. endo-1,4-beta-glucanase

Abstract

本发明涉及蛋白质工程领域，具体涉及一种高比活多功能纤维素酶突变体Nccle‑mut及其编码基因和应用。包含13个突变位点，分别为E18T，S20G，N25T，S35A，M80F，I91S，S122F，W127F、R150K、N158D、K162N、F165W、G208V以及V212A。突变体Nccle‑mut对羧甲基纤维素钠、葡聚糖以及木聚糖的相对酶活分别是原始酶Nccle的1.3倍，1.5倍和1.8倍。

Description

一种高比活多功能纤维素酶突变体Nccle-mut及其编码基因 和应用
技术领域
本发明涉及蛋白质工程领域，具体涉及一种高比活多功能纤维素酶突变体Nccle-mut及其编码基因和应用。
背景技术
纤维素和半纤维素作为世界上最丰富的可再生资源，一直受到人们的关注，成为生物质转化为简单糖发酵生产乙醇的关键。
纤维素酶主要包括外切纤维素酶、内切葡聚糖酶以及葡萄糖糖苷酶，半纤维素酶主要以木聚糖酶、甘露聚糖酶为主。此外自然界还存在着多功能纤维素酶，该酶不仅具有纤维素酶活性，同时还具有葡聚糖酶、木聚糖酶活性，因此多功能纤维素酶具有很好的应用潜力。
Nccle是一种多功能纤维素酶，该酶对羧甲基纤维素钠、木聚糖以及葡聚糖均具有水解活性，具有广泛的应用潜力。但是Nccle对羧甲基纤维素钠、木聚糖以及葡聚糖的水解比活力低，限制了其产业化应用。
发明内容
本发明对多功能纤维素酶Nccle进行分子改造，从而获得了高比活的多功能纤维素酶突变体Nccle-mut，降低了其生产成本，为多功能纤维素酶Nccle的产业化应用奠定基础。
本发明的目的是提供高比活多功能纤维素酶突变体Nccle-mut。
本发明的再一目的是提供高比活多功能纤维素酶突变体Nccle-mut的编码基因。
多功能纤维素酶Nccle-mut的核苷酸序列和氨基酸序列分别如SEQ ID NO.1和SEQID NO.3所示。
根据本发明的具体实施方式，采用易错PCR技术对SEQ ID NO.1所示的多功能纤维素酶Nccle的核苷酸序列进行改造，从而获得一系列突变体。经过筛选获得了14个有效的突变体，这14个有效单突变体分别为：E18T，S20G，N25T，S35A，M80F，I91S，S122F，W127F、R150K、N158D、K162N、F165W、G208V以及V212A。将易错PCR所获得的有效单突变体，进行叠加组合，最终获得一种高比活的多功能纤维素酶突变体Nccle-mut。突
根据本发明的纤维素酶突变体Nccle-mut共包含13个突变位点，分别为E18T，S20G，N25T，S35A，M80F，I91S，S122F，W127F、R150K、N158D、K162N、F165W、G208V以及V212A。突变体Nccle-mut对羧甲基纤维素钠、葡聚糖以及木聚糖的相对酶活分别是原始酶Nccle的1.3倍，1.5倍和1.8倍。突变体Nccle-mut的核苷酸序列如SEQ ID NO.2所示，氨基酸序列如SEQ ID NO.4所示。
本发明通过易错PCR技术和组合突变技术得到了一种高比活的多功能纤维素酶突变体Nccle-mut，为多功能纤维素酶Nccle的产业化应用奠定基础。
附图说明
图1显示原始多功能纤维素酶Nccle和突变体Nccle-mut的最适反应pH。
图2显示原始多功能纤维素酶Nccle和突变体Nccle-mut的pH稳定性。
图3显示原始多功能纤维素酶Nccle和突变体Nccle-mut的最适反应温度。
图4显示原始多功能纤维素酶Nccle和突变体Nccle-mut的热稳定性。
具体实施方式
以下实施例中未作具体说明的分子生物学实验方法，均参照《分子克隆实验指南》(第三版)J.萨姆布鲁克一书中所列的具体方法进行，或者按照试剂盒和产品说明书进行；所述试剂和生物材料，如无特殊说明，均可从商业途径获得。
实验材料和试剂：
1、菌株与载体
大肠杆菌菌株Topl0、毕赤酵母X33、载体pGAPzαA、Zeocin购自Invitrogen公司。
2、酶与试剂盒
Q5高保真Taq酶MIX购自NEB公司，质粒提取，胶纯化，限制性内切酶、试剂盒购自上海生工公司。
3、培养基
大肠杆菌培养基为LB(1％蛋白胨，0.5％酵母提取物，1％NaCl，pH7.0)。LBZ为LB培养基加25ug/mL Zeocin。
酵母培养基为YPD(1％酵母提取物，2％蛋白胨，2％葡萄糖)。酵母筛选培养基为YPDZ(YPD+100mg/L zeocin)。
实施例1、多功能纤维素酶Nccle基因克隆
根据已报道多功能纤维素酶Nccle的序列(Genebank:ORY82289)，直接合成Nccle基因，Nccle的基因序列如SEQ ID NO.1所示。根据合成基因的序列设计两条引物(R：5'-aggaattcGAATTTCAATCTTTCTGTTCT-3'和F：5'-attctagaTT AGTCACCAATATAAACCTT-3')用于扩增多功能纤维素酶Nccle基因。将扩增的PCR产物纯化回收，连接到表达载体pPGAPzαA-Nccle。
实施例2、易错PCR
以上述pPGAPzαA-Nccle为模板，进行易错PCR随机突变扩增，具体地扩增方法是：
第一轮扩增：以载体启动子引物AOX5-F和AOX3-R为引物进行PCR扩增，反应体系如下：
Buffer taq(10X) 10uL
易错dNTPs 10uL
引物1 1uL
引物2 1uL
pGAPzαA-Basamy 1uL
MgCl2 22uL
MnSO4 4uL
Taq酶 1uL
加水至 100uL
反应程序如下：
回收第一轮PCR产物，取1μL稀释50-100倍用作第二轮PCR的模板；第二，第三轮易错PCR以多功能纤维素酶Nccle基因特异性引物R及F替代引物AOX5-F和AOX3-R为反应引物，重复PCR反应。取第二、三轮的产物用XbaI和EcoRI进行双酶切，连接至pGAPzαA载体上的EcoRI和XbaI位点之间。连接产物转化X33，在YPDZ平板培养筛选突变菌株。将在YPDZ平板上长出来的菌接入YPD进行培养，培养36小时后，加入1％的甲醇进行诱导，诱导24小时后进行酶活测定(筛选阶段只进行纤维素酶活性的测定，纤维素酶活性测定参照轻工标准QB/T2583-2003进行)。
经过筛选获得了14个有效单突变体分别为E18T，S20G，N25T，S35A，M80F，I91S，S122F，W127F、R150K、N158D、K162N、F165W、G208V以及V212A。这14个单突变体的相对比活如表1所示。由表1可知，这14个有效的单突变体，均能一定程度的提高多功能纤维素酶Nccle的纤维素酶比活(6％至20％之间)。
表1 原始多功能纤维素酶Nccle和单突变体相对比活
编号 相对比活(％)
原始多功能纤维素酶Nccle 100
E18T 110
S20G 115
N25T 108
S35A 113
M80F 114
I91S 119
S122F 108
W127F 106
R150K 111
N158D 120
K162N 109
F165W 106
G208V 107
V212A 106
实施例3、组合突变
通过定点突变的方式，将所有单突变体组合到一起得到一种高比活的多功能纤维素酶突变体Nccle-mut，Nccle-mut包含的突变位点为E18T，S20G，N25T，S35A，M80F，I91S，S122F，W127F、R150K、N158D、K162N、F165W、G208V和V212A。分别测定突变体Nccle-mut的纤维素酶、葡聚糖酶以及木聚糖酶活性，由表2可知，相对于原始多功能纤维素酶Nccle，突变体Nccle-mut的纤维素酶、葡聚糖酶以及木聚糖酶的比活分别提高30％、50％和80％。
表2 原始多功能纤维素酶Nccle和突变体Nccle-mut对不同底物相对比活
实施例4、原始多功能纤维素酶Nccle和突变体Nccle-mut的最适反应pH
以羧甲基纤维素钠为底物，测定原始多功能纤维素酶Nccle和突变体Nccle-mut的最适反应pH。Nccle和突变体Nccle-mut的最适反应pH如图1所示。由图1可知，Nccle和突变体Nccle-mut，均为5.0。
实施例5、原始多功能纤维素酶Nccle和突变体Nccle-mut的pH稳定性
将多功能纤维素酶Nccle和突变体Nccle-mut分别在pH4-9条件下室温处理4小时，然后以羧甲基纤维素钠为底物测定酶活，结果如图2所示。由图2可知Nccle和突变体Nccle-mut的pH稳定性基本一样。
实施例6、原始多功能纤维素酶Nccle和突变体Nccle-mut的最适反应温度
以羧甲基纤维素钠为底物，测定原始多功能纤维素酶Nccle和突变体Nccle-mut的最适反应温度，结果如图3所示。由图3可知，原始多功能纤维素酶Nccle的最适反应温度为70℃，而突变体Nccle-mut的最适反应温度为60℃。
实施例7、原始多功能纤维素酶Nccle和突变体Nccle-mut的热稳定性
将多功能纤维素酶Nccle和突变体Nccle-mut分别在50℃-90℃条件下水浴处理30分钟，然后以羧甲基纤维素钠为底物测定酶活，结果如图4所示。由图4可知，原始多功能纤维素酶Nccle的热稳定性要好于突变体Nccle-mut，在80℃和90℃条件下，原始多功能纤维素酶Nccle的酶活保留率分别为60％和50％，而突变体Nccle-mut的保留率分别为52％和38％。
序列表
<110> 刘丹妮
<120> 一种高比活多功能纤维素酶突变体Nccle-mut及其编码基因和应用
<160> 4
<170> SIPOSequenceListing 1.0
<210> 1
<211> 684
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 1
gaatttcaat ctttctgttc ttctgcttcc cactccggtc agtctgtcaa ggaaacctct 60
aacaaggtcg gtaacatcgg tggagttgga tacgagttgt ggtctgactc cggtaacaac 120
tctgccacct tctactctga cggttccttc tcctgcacct ttcagaacgc cggtgactac 180
ttgtgccgtt ctggattgtc cttcgactcc accaaaaccc catcccaaat cggtcgttgg 240
aaggccgact ttaagttggt taaacaaaac atctccaacg ttggttactc ctacgtcggt 300
gtttacggtt ggaccagatc ccctttggtc gagtactaca tcgtcgacaa ctggttgtct 360
ccatctccac caggtgactg ggttggtaac aagaagcacg gttccttcac cattgacgga 420
gctcagtaca ccgtttacga gaacaccaga accggtccat ccattgatgg taacactact 480
tttaaacaat acttctccat cagacagcag gccagagact gtggtaccat tgacatctcc 540
gcccactttg accagtggga gaagttgggt atgaccatgg gtaagttgca cgaggctaag 600
gtcttgggtg aggctggtaa cggtaacggt ggtgtttctg gtaccgccga cttcccatac 660
gctaaggttt atattggtga ctaa 684
<210> 2
<211> 684
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 2
gaatttcaat ctttctgttc ttctgcttcc cactccggtc agtctgtcaa ggttaccggt 60
aacaaggtcg gtaccatcgg tggagttgga tacgagttgt gggctgactc cggtaacaac 120
tctgccacct tctactctga cggttccttc tcctgcacct ttcagaacgc cggtgactac 180
ttgtgccgtt ctggattgtc cttcgactcc accaaaaccc catcccaaat cggtcgtttc 240
aaggccgact ttaagttggt taaacaaaac tcttccaacg ttggttactc ctacgtcggt 300
gtttacggtt ggaccagatc ccctttggtc gagtactaca tcgtcgacaa ctggttgtct 360
ccattcccac caggtgactt cgttggtaac aagaagcacg gttccttcac cattgacgga 420
gctcagtaca ccgtttacga gaacaccaaa accggtccat ccattgatgg tgacactact 480
tttaaccaat actggtccat cagacagcag gccagagact gtggtaccat tgacatctcc 540
gcccactttg accagtggga gaagttgggt atgaccatgg gtaagttgca cgaggctaag 600
gtcttgggtg aggctggtaa cgttaacggt ggtgcttctg gtaccgccga cttcccatac 660
gctaaggttt atattggtga ctaa 684
<210> 3
<211> 227
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 3
Glu Phe Gln Ser Phe Cys Ser Ser Ala Ser His Ser Gly Gln Ser Val
1 5 10 15
Lys Glu Thr Ser Asn Lys Val Gly Asn Ile Gly Gly Val Gly Tyr Glu
20 25 30
Leu Trp Ser Asp Ser Gly Asn Asn Ser Ala Thr Phe Tyr Ser Asp Gly
35 40 45
Ser Phe Ser Cys Thr Phe Gln Asn Ala Gly Asp Tyr Leu Cys Arg Ser
50 55 60
Gly Leu Ser Phe Asp Ser Thr Lys Thr Pro Ser Gln Ile Gly Arg Met
65 70 75 80
Lys Ala Asp Phe Lys Leu Val Lys Gln Asn Ile Ser Asn Val Gly Tyr
85 90 95
Ser Tyr Val Gly Val Tyr Gly Trp Thr Arg Ser Pro Leu Val Glu Tyr
100 105 110
Tyr Ile Val Asp Asn Trp Leu Ser Pro Ser Pro Pro Gly Asp Trp Val
115 120 125
Gly Asn Lys Lys His Gly Ser Phe Thr Ile Asp Gly Ala Gln Tyr Thr
130 135 140
Val Tyr Glu Asn Thr Arg Thr Gly Pro Ser Ile Asp Gly Asn Thr Thr
145 150 155 160
Phe Lys Gln Tyr Phe Ser Ile Arg Gln Gln Ala Arg Asp Cys Gly Thr
165 170 175
Ile Asp Ile Ser Ala His Phe Asp Gln Trp Glu Lys Leu Gly Met Thr
180 185 190
Met Gly Lys Leu His Glu Ala Lys Val Leu Gly Glu Ala Gly Asn Gly
195 200 205
Asn Gly Gly Val Ser Gly Thr Ala Asp Phe Pro Tyr Ala Lys Val Tyr
210 215 220
Ile Gly Asp
225
<210> 4
<211> 227
<212> PRT
<213> 人工序列(Artificial Sequence)
<400> 4
Glu Phe Gln Ser Phe Cys Ser Ser Ala Ser His Ser Gly Gln Ser Val
1 5 10 15
Lys Val Thr Gly Asn Lys Val Gly Thr Ile Gly Gly Val Gly Tyr Glu
20 25 30
Leu Trp Ala Asp Ser Gly Asn Asn Ser Ala Thr Phe Tyr Ser Asp Gly
35 40 45
Ser Phe Ser Cys Thr Phe Gln Asn Ala Gly Asp Tyr Leu Cys Arg Ser
50 55 60
Gly Leu Ser Phe Asp Ser Thr Lys Thr Pro Ser Gln Ile Gly Arg Phe
65 70 75 80
Lys Ala Asp Phe Lys Leu Val Lys Gln Asn Ser Ser Asn Val Gly Tyr
85 90 95
Ser Tyr Val Gly Val Tyr Gly Trp Thr Arg Ser Pro Leu Val Glu Tyr
100 105 110
Tyr Ile Val Asp Asn Trp Leu Ser Pro Phe Pro Pro Gly Asp Phe Val
115 120 125
Gly Asn Lys Lys His Gly Ser Phe Thr Ile Asp Gly Ala Gln Tyr Thr
130 135 140
Val Tyr Glu Asn Thr Lys Thr Gly Pro Ser Ile Asp Gly Asp Thr Thr
145 150 155 160
Phe Asn Gln Tyr Trp Ser Ile Arg Gln Gln Ala Arg Asp Cys Gly Thr
165 170 175
Ile Asp Ile Ser Ala His Phe Asp Gln Trp Glu Lys Leu Gly Met Thr
180 185 190
Met Gly Lys Leu His Glu Ala Lys Val Leu Gly Glu Ala Gly Asn Val
195 200 205
Asn Gly Gly Ala Ser Gly Thr Ala Asp Phe Pro Tyr Ala Lys Val Tyr
210 215 220
Ile Gly Asp
225

Claims (7)

1.一种高比活的多功能纤维素酶突变体Nccle-mut，其特征在于，所述突变体的氨基酸序列为SEQ ID NO.3所示的氨基酸序列的18位由E突变为T；20位由S突变为G；25位由N突变为T；35位由S突变为A；80位由M突变为F；91位由I突变为S；122位由S突变为F；127位由W突变为F；150位由R突变为K；158位由N突变为K；162位由K突变为N；165位由F突变为W；208位由G突变为V；和212为由V突变为A。
2.编码权利要求1所述的高比活的多功能纤维素酶突变体Nccle-mut的基因。
3.根据权利要求2所述的基因，其特征在于，所述基因的核苷酸序列如SEQ ID NO.2所示。
4.包含权利压强2所述基因的重组载体。
5.包含权利要求/2所述基因的宿主细胞。
6.一种制备高比活的多功能纤维素酶的方法，其特征在于，所述方法包括将权利要求4所述的重组载体表达宿主细胞，表达并分离高比活的多功能纤维素酶的步骤。
7.权利要求1所述的高比活的多功能纤维素酶突变体Nccle-mut的应用。
CN201810085865.0A 2018-01-29 2018-01-29 一种高比活多功能纤维素酶突变体Nccle-mut及其编码基因和应用 Pending CN108103047A (zh)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201810085865.0A CN108103047A (zh) 2018-01-29 2018-01-29 一种高比活多功能纤维素酶突变体Nccle-mut及其编码基因和应用

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201810085865.0A CN108103047A (zh) 2018-01-29 2018-01-29 一种高比活多功能纤维素酶突变体Nccle-mut及其编码基因和应用

Publications (1)

Publication Number Publication Date
CN108103047A true CN108103047A (zh) 2018-06-01

Family

ID=62221400

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201810085865.0A Pending CN108103047A (zh) 2018-01-29 2018-01-29 一种高比活多功能纤维素酶突变体Nccle-mut及其编码基因和应用

Country Status (1)

Country Link
CN (1) CN108103047A (zh)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20220195410A1 (en) * 2020-12-17 2022-06-23 Fornia Biosolutions, Inc. Xylanase Variants and Methods

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101210236A (zh) * 2006-12-30 2008-07-02 中国科学院上海生命科学研究院 多功能纤维素酶及其应用
CN104109660A (zh) * 2013-04-17 2014-10-22 东莞泛亚太生物科技有限公司 酶活性提高的木聚糖酶
CN104726434A (zh) * 2015-03-27 2015-06-24 云南师范大学 一种木聚糖酶XynRBM26及其编码基因
WO2017100429A1 (en) * 2015-12-09 2017-06-15 The Regents Of The University Of California Novel proteins from anaerobic fungi and uses thereof

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101210236A (zh) * 2006-12-30 2008-07-02 中国科学院上海生命科学研究院 多功能纤维素酶及其应用
CN104109660A (zh) * 2013-04-17 2014-10-22 东莞泛亚太生物科技有限公司 酶活性提高的木聚糖酶
CN104726434A (zh) * 2015-03-27 2015-06-24 云南师范大学 一种木聚糖酶XynRBM26及其编码基因
WO2017100429A1 (en) * 2015-12-09 2017-06-15 The Regents Of The University Of California Novel proteins from anaerobic fungi and uses thereof

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
HAITJEMA,C.H.等: "Neocallimastix californiae strain G1 LY90scaffold_9, whole genome shotgun sequence GenBank: MCOG01000009.1", 《GENBANK》 *
无: "LY90DRAFT_697603 - Endo-1,4-beta-xylanase - Neocallimastix californiae", 《EMBL》 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20220195410A1 (en) * 2020-12-17 2022-06-23 Fornia Biosolutions, Inc. Xylanase Variants and Methods

Similar Documents

Publication Publication Date Title
US10947568B2 (en) Limiting yeast-produced trehalose in fermentation
US11332728B2 (en) Yeast strains for the expression and secretion of heterologous proteins at high temperatures
RU2007124552A (ru) Глюкоамилаза trichoderma reesei и ее гомологи
CN107012130A (zh) 一种葡萄糖氧化酶突变体及其编码基因和应用
CN111334494A (zh) 新型高稳定性碱性蛋白酶突变体的筛选
CN106591271A (zh) 一株酶活和温度稳定性提高的精氨酸脱亚胺酶突变体及其应用
CN111518792B (zh) 一种草酸青霉16的β-葡萄糖苷酶突变体TLE及应用
CN106754825B (zh) 提高比活的α-淀粉酶BaAmy突变体及其编码基因和应用
WO2005105991A1 (ja) 放線菌由来のａｍｐデアミナーゼ、及びその使用
CN108103047A (zh) 一种高比活多功能纤维素酶突变体Nccle-mut及其编码基因和应用
EP2897975A1 (en) Modified fungal cell
CN110713999B (zh) 提高比活的α-淀粉酶突变体BasAmy-3及其编码基因和应用
KR101235422B1 (ko) 백색부후균 유래의 엔도-1,4-베타 자일라나제 ａ 대량생산을 위한 재조합 발현벡터 및 이를 이용한 엔도-1,4-베타 자일라나제 ａ 대량생산방법
CN107022536B (zh) 高催化效率的纤维素酶突变体及其基因和应用
WO2016175202A1 (ja) 耐熱性を有するキシラナーゼ
US9650625B1 (en) Thermostable pectinase
CN112553182A (zh) 具提升活性的α-半乳糖苷酶
Zhang et al. Characterization, gene cloning and expression of new xylanase XYNB with high specific activity
KR20130027984A (ko) 활성이 증진된 변이 베타-글루코시다제 및 이를 이용한 바이오 에탄올의 제조방법
CN114457058B (zh) 一种饲用α淀粉酶的突变改良方法及应用
KR101218383B1 (ko) 열안정성 및 내산성을 가지는 엔도글루칸아제-디의 돌연변이체 및 그 제조방법
KR101235406B1 (ko) 백색부후균 유래의 엔도-1,4-베타 자일라나제 ａ 대량생산을 위한 재조합 발현벡터 및 이를 이용한 엔도-1,4-베타 자일라나제 ａ 대량생산방법
KR101235528B1 (ko) 백색부후균 유래의 엔도-1,4-베타 자일라나제 ｃ대량생산을 위한 재조합 발현벡터 및 이를 이용한 엔도-1,4-베타 자일라나제 ｃ대량생산방법
Feng et al. Expression and characterization of a novel endo-1, 4-β-xylanase produced by Streptomyces griseorubens JSD-1 isolated from compost-treated soil
Feng et al. Insight into a novel β-1, 4-glucosidase from Streptomyces griseorubens JSD-1

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
RJ01 Rejection of invention patent application after publication
RJ01 Rejection of invention patent application after publication

Application publication date: 20180601