褐腐真菌产生的低分子化合物对纤维素的解聚作用研究

糙皮侧耳（Ｐｌｅｕｒｏｔｕｓｏｓｔｒｅａｔｕｓ）密粘褶菌（Ｇｌｏｅｏｐｈｙｌｌｕｍｔｒａｂｅｕｍ）洁丽香菇（Ｌｅｎｔｉｎｕｓｌｅｐｉｄｅｕｓ）等８株褐腐菌在滤纸上进行固体培养时,在培养初期,纤维素的聚合度均呈现大幅度下降,但不表现失重。培养过程中也未能检测出滤纸酶活力,只有少量内切葡聚糖酶活力。而且这８株菌都具有络合Ｆ３＾３＋和产生羟基自由基,ＯＨ的能力,由降解和解聚能力最强的密粘褶菌的胞外酶液在     

密粘褶菌Gloeophyllum trabeum胞外低分子量多肽的分离纯化及其对纤维素生物降解作用

通过HPLC高效液相层析由褐腐真菌中能强烈降解木质纤维素的代表菌株密粘褶菌Gloeo phyllumtrabeum的胞外培养液 ,分离纯化得到一低分子量的活性多肽组分 (Gt因子 ) .Gt因子具有较好热稳定性 ,在pH 2 5～ 6 5范围内保持稳定 .Gt因子分子量在 4 0 0 0左右 ,等电点pI 6 6 .Gt因子具有络合Fe3 + 的能力 ,且能够将Fe3 + 还原为Fe2 + .在O2 存在时 ,能以纤维素物质为电子供体形成羟基自由基HO·.利用循环伏安法 ,观察到Gt因子与纤维素底物之间的氧化还原过程 .研究表明 ,Gt因子极有可能在褐腐菌的纤维素降解初期起着重要的作用 .     

青海高原降解纤维素微生物的调查、分离、鉴定

从青海高原林区分离筛选 3 0 0余株分解纤维素的细菌及 3 1株降解纤维素的真菌。测定纤维素分解菌含量土样为 2 6× 1 0 5 g。对纤维素酶水解圈较大的 1 1株真菌 ,根据其滤纸酶活筛选出一株分离自互助北山森林的高产纤维素酶的真菌No 0 1 43菌株 ,根据其形态学及培养特征鉴定为康氏木霉 (TrichodermakoningiiQudem) ,该菌湿固体发酵物含滤纸酶活力(FPA)为 1 5u g。该菌无毒副作用 ,可用于饲料业     

青海高原降解纤维素微生物的调查、分离、鉴定*

从青海高原林区分离筛选 300余株分解纤维素的细菌及 31株降解纤维素的真菌。测定纤维素分解菌含量土样为 2.6×10⁵ g。对纤维素酶水解圈较大的11株真菌 ,根据其滤纸酶活筛选出一株分离自互助北山森林的高产纤维素酶的真菌No. 0143菌株 ,根据其形态学及培养特征鉴定为康氏木霉 (Trichodermakoningii Qudem) ,该菌湿固体发酵物含滤纸酶活力(FPA)为15u/g。该菌无毒副作用 ,可用于饲料业。     

地衣芽孢杆菌碱性蛋白酶的研究 Ⅰ.碱性蛋白酶高产菌的筛选及产酶条件初探

从成都佳丰食品厂等处采集的样品中平板分离初筛到124株碱性蛋白酶产生菌,进一步复筛出一株高产,且稳定的碱性蛋白酶产生菌株B．L．JF-ld,初步鉴定为地衣穿孢杆菌（Bacilluslicheniformis）。该菌的最适产酶条件为：培养基（％）为麦芽糖7．5,酵母膏3,NaCl0．5,K2HPO4·3H2O0．53,NaHPO4·2H2O0．03,Na2CO30．056,MnSO4l×10－4mol／L,pH8．7,通气量为（1：0．5）～（1：1）（v／v）,37℃发酵40h,酶活力单位高达7180U／ml。     

高效厌氧纤维素降解细菌的分离及酶特性研究

采用透明圈初筛和滤纸降解率复筛的方法从内蒙古绵羊瘤胃内容物中分离到高效厌氧纤维素降解细菌4株.通过形态学、生理生化反应、生态特性和遗传型的鉴定,所分离的4株菌WHQ、LYQ、LBG-1和NDF-3分别归为溶纤维丁酸弧菌(Butyrivibrio fibrisollvens)、黄色瘤胃球菌(Ruminococcus flavefaciens)、产琥珀酸丝状杆菌(Fibrobacter succinogenes)和解多糖梭菌(Clostridium polysaccharolyticum).测定了4株菌对滤纸的降解率,WHQ、LYQ、LBG-1和NDF-3的2周滤纸降解率分别为25.1%、14.3%、21.0%和20.6%.本研究同时对4株菌的滤纸酶活力、羧甲基纤维素酶活力和β-葡萄糖苷酶活力进行了测定.     

秸秆纤维素分解菌的酶活力测定

目的:测定秸秆纤维素分解菌的酶活力。方法:从土壤中分离出具有分解纤维素能力的菌株,采用刚果红染色法进行粗选,得到7株透明圈较大的菌株。将这7株菌株液体发酵培养6d,再分别用滤纸分解度观察、羧甲基纤维素酶活法(CMC)、滤纸酶活法(FPA)和天然纤维素酶活法测定其酶活力。结果:在7株菌株中,F-1、F-2、F-3、F-5的酶活力测定结果与其溶解圈的测定结果、滤纸分解结果基本相同。且天然纤维素酶活力高的菌株,其CMC酶活、FPA酶活也高,滤纸分解效果也比较明显。结论:CMC法、FPA法和天然纤维素酶活法适于测定秸秆纤维素分解菌的酶活力。     

密粘褶菌胞外低分子量多肽在纤维素降解中作用的研究

从褐腐真菌中能强烈降解纤维素的代表菌株密粘褶菌(Gloeophyllum trabeum)的胞外酶液中首次分离纯化得到一低分子量的活性多肽组分(称作Gt因子),此组分能在有O.2和Fe³⁺存在时产生羟基自由基HO·;对纤维素降解的研究表明,Gt因子不同于纤维素酶对纤维素的β1.4糖苷键的水解作用,而以HO·氧化的途径作用于纤维素,导致纤维素中氢键的断裂,降低纤维素的结晶度,使其暴露出更多的末端,从而有利于纤维素的进一步降解。     

根霉植酸酶的研究──Ⅰ.菌株的分离筛选及发酵条件研究

从21个土样和15株产酶菌株中筛选到1株产酶能力较强的菌株——根霉（Rhizopussp.）MR020。其优化培养基组成（％）：大米粉2.0,干酪素0．05,（NH4）2SO41.0,MgSO4·7H2O0.05,FeSO4·7H2O0.01,pH5.0。其振荡培养条件：培养基装最25ml／250ml三角瓶,用培养8天、浓度为1×107个／ml的孢子悬液接种1ml,于30℃,150r／min振荡培养108h产酶量最高达4000u/ml。     

中温（37℃）纤维素分解菌的筛选及混合培养研究

目的:筛选纤维素分解菌,构建复合微生物菌系进行混合培养,获得降解纤维素能力强的复合菌系.方法:从高温阶段堆肥样品、腐熟肥料、牛粪和土壤中筛选出能较好降解纤维素的菌株,进行单独与混合发酵培养,测其酶活.结果:获得降解纤维素能力较强的细菌3株、霉菌4株、放线菌2株.按不同的接种比例构建了4组复合微生物菌系,4个组合的滤纸失重率分别达到41.52%、44.94%、41.82%、37.11%,液体发酵的平均CMC酶活分别为624U/g、988U/g、769U/g、1041U/g,固体发酵的平均CMC酶活分别为4240U/g、5289U/g、4807U/g、5344U/g.结论:综合分析复合菌系滤纸条降解能力、CMC酶活、FAP酶活表明,组合二分解纤维素能力明显强于其他几个组合.     

褐腐真菌产生的羟基自由基HO˙对纤维素作用的研究*

由褐腐真菌的典型菌株——密粘褶菌Gloeophyllum trabeum的胞外培养液中分离纯化得到一能还原Fe3+,产生羟基自由基HO˙的多肽组分(称作Gt因子)。 采用HO˙特异性的抑制剂硫脲,对Gt因子产生的HO˙在纤维素降解中的作用进行了对照研究,结果表明Gt因子及其产生的HO˙在纤维素降解中起着重要的作用,为褐腐菌HO˙氧化降解纤维素机制假说的确立提供了一些依据。     

Mechanisms of wood degradation by brown-rot fungi: chelator-mediated cellulose degradation and binding of iron by cellulose

Iron, hydrogen peroxide, biochelators and oxalate are believed to play important roles in cellulose degradation by brown-rot fungi. The effect of these compounds in an 'enhanced' Fenton system on alpha-cellulose degradation was investigated specifically in regard to molecular weight distribution and cellulose-iron affinity. This study shows that the degradative ability of an ultrafiltered low molecular weight preparation of chelating compounds isolated from the brown-rot fungus Gloeophyllum trabeum (termed 'Gt chelator') increased with increasing Gt chelator concentration when the FeIII to Gt chelator ratio was greater than about 30:1. When this ratio was less than 30:1, increasing Gt chelator concentration did not accelerate cellulose degradation. In excess hydrogen peroxide, cellulose degradation increased and then decreased with increasing iron concentration when FeIII was present in excess of the Gt chelator. The critical ratio of FeIII to Gt chelator varied depending on the concentration of hydrogen peroxide in the system. Increasing iron concentration above a critical iron:chelator ratio inhibited cellulose degradation. The optimum pH for cellulose degradation mediated by Gt chelator was around 4.0. A comparison of the effects of 2,3-DHBA (a chelator that reduces iron similarly to Gt chelator) and Gt chelator with respect to cellulose degradation demonstrated the same pattern of cellulose degradation. Cellulose-iron affinity studies were conducted at three pH levels (3.6, 3.8, 4.1), and the binding constants for cellulose-FeIII, cellulose-FeII and cellulose-FeIII in the presence of Gt chelator were calculated. The binding constants for cellulose-FeIII at all three pH levels were much higher than those for cellulose-FeII, and the binding constants for cellulose-FeIII in the presence of Gt chelator were very close to those for cellulose-FeII. This is probably the result of FeIII reduction to FeII by Gt chelator and suggests that chelators from the fungus may be able to sequester iron from cellulose and reduce it in near proximity to the cellulose and thereby better promote depolymerization. The free radical generating system described has potential for use in a variety of industrial processing and pollution control applications.     

新分离菌Streptomyces产小分子CMC液化酶的研究

新分离纤维素分解菌经鉴定为链霉菌属中的新种,暂定名为ＳｔｒｅｐｔｏｍｙｃｅｓｓｐＬＸ,革兰氏阳性,产气生孢子,好氧生长,最适生长温度和ＰＨ为３０℃和７．２。该菌能够完全降解纤维素且不产生还原糖,并分泌一种分子量为９．２ｋｕ的液化性质的内切纤维素液化酶,亦即ＣＭＣ液化酶,该酶在裂解滤纸为短纤维的过程中既没有还原糖生成,也没有失重现象发生,而且纤维素的聚合度也几乎没有明显变化,推测可能是一种能打开氢键的小分子解链酶。     

棉秸秆降解高温菌株的筛选及产酶分析

从新疆地区分离具有降解棉秸秆纤维素功能的菌株,得到4株耐高温真菌(50°C)。纤维素酶学性质分析表明,该4株菌的纤维素酶具有良好的耐酸性(最适pH为4.5)和耐高温性(最高达60°C)。以羧甲基纤维素钠(CMC-Na)、微结晶纤维素、棉花、滤纸、淀粉、果胶为底物测定酶活力,滤纸酶活力(FPA)最高达2.63 U/mL、淀粉酶活力最高达6.17 U/mL、果胶酶活力最高达5.86 U/mL。4株真菌酶学特性分析表明,该系列菌株在秸秆生物质利用方面有很大的应用潜力。     

Function and mechanism of a low-molecular-weight peptide produced by Gloeophyllum trabeum in biodegradation of cellulose

A special low-molecular-weight peptide named Gt factor, was isolated and purified via HPLC from the culture extract of the brown-rot fungus Gloeophyllum trabeum. It had high-affinity Fe(3+)-chelating ability and could reduce Fe(3+) to Fe(2+). In the presence of O(2), it could produce hydroxyl radicals HO*. The effects of Gt factor on cellulose degradation suggested that Gt factor could disrupt inter- and intra- hydrogen bonds in cellulose chains by a HO*-involved mechanism. This resulted in depolymerization of cellulose chains, which produced more reducing and non-reducing ends, thus making cellulose accessible for further degradation. This pathway was quite different from the hydrolytic processes driven by cellulases, and Gt factor might play an important role in the early stage of cellulose depolymerization by brown-rot fungi.     

甘蔗渣纤维素降解菌株的筛选与鉴定

甘蔗渣是制糖工业的主要副产物。筛选甘蔗渣纤维素降解菌株对甘蔗渣乙醇产业具有重要的意义。以甘蔗渣为原料,通过分离和纯化得到14株菌株,对其进行纤维素刚果红平板染色实验和滤纸崩解实验,最终获得3株可以生产纤维素酶的菌株02-2-2、21-1-2和40-1-1。酶活性测定结果表明,菌株40-1-1的酶活力在培养3 d后达到最高,为27.26 U/mg。通过形态学和分子生物学鉴定,菌株02-2-2为枝顶孢属(Acremonium sp.),菌株21-1-2和40-1-1为光滑短梗霉属(Acrophialophora sp.)。研究筛选的菌株将为开展甘蔗渣纤维素降解利用提供参考。     

Cellulase production from the corn stover fraction based on the organ and tissue

In this study, the biodegradation of fractionated corn stover in solid-state fermentation by Trichoderma reesei YG3 was investigated. Fractions of miscellaneous cells (MC) and fasciculi (FC) tissue from leaf, shell or core were separated using the combined pretreatment method of carding classification after steam explosion. The highest enzyme activities including the filter paper activity, endoglucanase, exoglucanase and β-glucosidase activities, weight loss rate of dry material and biodegradation rate were all observed in the MC tissue fraction from the leaf, which was more nutritious, while lowest activity was observed in the FC tissue fraction from the shell. The maximum filter paper activity and weight loss rate of the dry material were 4.56 and 1.89 times the minimum and the cellulose and hemicellulose biodegradation rates were 51.22 and 39.38% versus 23.85 and 26.51%, respectively. These variances maybe attributed to the heterogeneity of the component in the fractions. A higher weight loss rate corresponded to higher enzymatic activities, whereas cellulose biodegradation was not proportional to cellulase activities. Hemicellulose biodegradation was much slower than cellulose degradation. Here, we demonstrated the importance of fractionation in component biodegradation and utilization of straw.     

Culm Brittleness of Barley (Hordeum vulgare L.) Mutants Is Caused by Smaller Number of Cellulose Molecules in Cell Wall

The physicochemical nature of the cell wall was determined in the fourth internode of three isogenic brittle mutants of barley (Hordeum vulgare L.) and corresponding nonbrittle strains. Cellulose contents of the brittle culms were 17.5 to 20.3% of those of corresponding nonbrittle strains. No major difference was found in lignin and noncellulose components (except glucose) between brittle and nonbrittle strains. Maximum bending stresses of brittle culms were 38.0 to 54.2% of those of corresponding nonbrittle strains. The degree of polymerization of cellulose, measured by viscometry, was similar between the brittle and the nonbrittle strains. Mole number of cellulose molecules in a unit length of brittle culms, calculated by dividing cellulose mass by molecular weight, was 7.7 to 17.3% of those of the nonbrittle strains. These results indicate that brittleness of mutant culms is due to fewer numbers of cellulose molecules in the cell walls.     

象白蚁肠道中一个纤维素降解菌群的分离和特性研究

利用滤纸培养基从象白蚁(Nasutitermes sp.)肠道中分离出一个具有纤维素降解能力,能够降解滤纸的混合菌群。在起始pH 6.5,37℃培养条件下培养6d可得到最高的纤维素酶(CMCase和FPase)活性。在优化条件下,混合菌群的滤纸降解率在第15d达到最大值66.3%,显示出较高的滤纸降解效率。酶谱活性染色分析显示,混合菌群在以滤纸为唯一碳源的生长过程中至少表达了8种内切葡聚糖酶和4种木聚糖酶。扫描电镜观察到该混合菌群包含短杆状和球形两种形态的细菌。基于16SrRNA基因的系统发育分析表明,该混合菌群中至少存在两种细菌,分别属于沙雷氏菌属(Serratia)和类芽胞杆菌属(Paenibacillus)。这两种细菌协同降解纤维素的机制值得进一步深入研究。