4种菌根真菌对五氯酚耐受性及其生理基础研究

通过纯培养的方法，研究了4种外生菌根真菌——美味牛肝菌(Boletus edulis)、褐环乳牛肝菌(Suillus luteus)、丝膜菌、(Cortinarrus russus)和厚环粘盖乳牛肝菌(Suillus grevillei)的生长效应、对五氯酚的耐受性，并采用点试方法对真菌氧化还原酶的产生情况进行了检测。结果表明，4种外生菌根真菌在MM培养基上的生长速率显著高于MMN培养基。4种外生菌根真菌对五氯酚均有一定的耐受性，厚环粘盖乳牛肝菌(Suillus grevillei)耐受性最高，其生长抑制率显著低于其它真菌。酶测试结果表明，厚环粘盖乳牛肝菌是4种真菌中氧化还原酶活性较强且酶种类多样的菌株，它表现出很高的过氧化物酶、多酚氧化酶和漆酶活性，是一种具有高效降解芳环结构污染物潜力的菌根真菌。4种真菌在生物量和“酸化效应”方面差异较大。厚环粘盖乳牛肝菌(Suillus grevillei)对五氯酚的高耐受性可能与其产酶特性和“酸化效应”有关。     

农业废弃物堆腐过程中氧化还原酶活性的变化

【目的】从酶学角度揭示好氧发酵的生物机理。【方法】选择在目前我国数量多、分布广的有机固体废弃物鸡粪和小麦秸秆及花椒籽粕为主要材料,采用堆腐装置,以未添加复合微生物菌剂作为对照(CK),研究复合微生物菌剂(加菌剂处理)对农业废弃物静态高温堆腐过程中氧化还原酶(过氧化氢酶、脱氢酶和多酚氧化酶)活性的影响。【结果】加菌剂处理和CK的过氧化氢酶活性分别在堆腐3和4 d达到峰值(1 705和1 697 mL/g);加菌剂处理和CK的脱氢酶活性分别在堆腐10和12 d达最大值(8.39和6.57μL/g);加菌剂处理和CK的多酚氧化酶活性分别在堆腐18和32 d达峰值(7.33和6.65 mg/g)。从堆腐的整个过程看,随着堆腐时间的延长,加菌剂处理的过氧化氢酶、脱氢酶和多酚氧化酶活性上升较快,而且这3种酶活性的最大值均比CK高。【结论】添加复合微生物菌剂可以提高堆腐过程中氧化还原酶活性,使酶活性峰值出现的时间提前,对堆腐过程中的物质分解具有促进作用。     

铜绿微囊藻对亚硝态氮的利用

通过室内培养,研究了不同亚硝态氮浓度对铜绿微囊藻（Microcystis aeruginosa）生长的影响和藻对亚硝态氮的利用,实验分析了水体中亚硝态氮、硝态氮和铵态氮浓度的变化,测定了铜绿微囊藻的生长曲线、藻细胞内亚硝态氮含量和藻亚硝酸氧化酶（NOR）。结果显示,在10mgNO^-2-N·L^-1的处理组中,培养基中亚硝态氮和硝态氮浓度同时减少,说明铜绿微囊藻可以同时利用亚硝态氮和硝态氮;在20和30mgNO^-2-N·L^-1的处理组中,随着藻的生长培养基中亚硝态氮的浓度减少,硝态氮浓度增加,而且电泳实验显示此培养条件下铜绿微囊藻能产生亚硝酸氧化酶,表明培养基中的亚硝态氮被亚硝酸氧化酶氧化为硝态氮。本实验也表明高浓度的亚硝态氮（大于10mgNO^-2-N·L^-1）能够抑制藻的生长。     

重金属锌对猪粪堆肥过程中氧化还原类酶活性的影响

以猪粪和秸秆为主要试验材料,添加不同浓度重金属Zn,采取发酵罐处理方法,在好氧高温条件下研究了重金属Zn对猪粪堆肥过程中多酚氧化酶、脱氢酶活性的变化,以及堆腐过程堆体温度、堆料pH值、胡敏酸E4/E6值的变化。结果表明：（1）低量重金属Zn处理（L）较不添加重金属Zn（CK）和添加高量重金属Zn（H）堆料升温快、温度高、高温持续时间长。（2）重金属Zn的加入对堆料的pH值影响不大,不是影响堆肥进程的直接原因。（3）H处理在整个堆肥过程中E4/E6值均高于L和CK,表明高浓度Zn处理抑制腐殖质的缩合和芳构化。（4）L处理的多酚氧化酶活性大多数时间高于H处理的活性,说明低量重金属Zn更好地促进了木质素的降解及其产物的转化。（5）从整个堆肥过程来看,3个不同处理的脱氢酶活性表现出一定的不稳定性,可能是重金属对脱氢酶活性有抑制作用的同时发生＂抗性酶活性＂现象。     

人工湿地氧化还原特征及其与微生物活性相关性

在水力负荷为800 mm/d的运行条件下,研究了复合垂直流人工湿地中试系统不同功能层面的氧化还原特征、好氧微生物数量、基质的耗氧速率及氧化还原酶类活性的分布特点。结果表明:各功能层面的氧化还原电位、好氧微生物数量、耗氧速率表层最高,并且随着基质深度的增加逐渐降低。除过氧化物酶外,氧化酶活性表现为表层高于中下层;而硝酸盐还原酶、亚硝酸盐还原酶等活性变化趋势则相反。同时,氧化还原电位与基质中细菌、真菌、放线菌的数量、耗氧速率等存在显著正相关关系;氧化还原电位与多酚氧化酶、过氧化氢酶、脱氢酶等氧化酶活性也存在显著正相关,而与下行流池的硝酸盐还原酶、亚硝酸盐还原酶等还原酶类存在显著负相关关系。     

Screening of proteins interacting with activation domain of macrophage migration-inhibitory factor

AIM: To screen the proteins that interact with the catalytic thiol-protein oxidoreductase (TPOR) domain of macrophage migration-inhibitory factor (MIF). METHODS: Yeast two-hybrid system was used to study the proteins that interacted with MIF. The bait vector pBTM116-MIF was constructed and transfected into L40 yeast strain. L40 competent cells expressing the TPOR domain were prepared and used to screen the proteins interacting with the TPOR domain of MIF in a human osteosarcoma cDNA library. The proteins interacting with TPOR domain were identified by HIS3 reporter gene and β-galactosidase assay. The techniques of co-immunoprecipitation and immunofluorescence were also used to verify the proteins interacting with TPOR domain. RESULTS: Wild-type pBTM116-MIF and pACT2-TXNL2 (thioredoxin-like 2 protein) were constructed and co-transfected into L40 yeast strain. The activation of HIS3 reporter gene and the positive result of β-galactosidase assay indicated that TXNL2 was the candidate protein that interacted with the catalytic TPOR domain of MIF. The vector of pcDNA3.1-Myc-TXNL2 was constructed and transfected into MCF-7 cell line. The result of co-immunoprecipitation assay showed that the catalytic TPOR domain of MIF co-immunoprecipitated with Myc-TXNL2 protein. The result of immunofluorescence assay demonstrated that the catalytic TPOR domain of MIF and TXNL2 co-localized in the cytoplasm of the cells. CONCLUSION: TXNL2 is the protein that interacts with the catalyzed TPOR domain of MIF.     

克雷伯氏菌1,3-丙二醇氧化还原酶基因在大肠杆菌中的克隆与表达

以基因组DNA为模板,利用PCR技术从克雷伯氏菌(Klebsiella pneumoniae)中扩增得到含有1,3-丙二醇氧化还原酶基因(dhaT)的DNA片段,将其定向连接到克隆质粒pGEM-3xf上,得到重组质粒pGEM-dhaT。将此重组质粒转化到受体菌Escherichia coli DH5α中,通过蓝白斑鉴定挑选出阳性菌株。DNA序列分析表明其基因全长为1185bp。将该片段插入表达载体pSE380,构建成重组子pSE-dhaT,并在E．coli JM109中获得表达,经SDS-PAGE检测,表达产物分子质量与天然纯1,3-丙二醇氧化还原酶(DHAT)相同,约为43ku。     

1,3-丙二醇氧化还原酶基因的克隆与序列分析

以肺炎克雷伯氏菌的基因组DNA为模板,通过PCR扩增得到了目的基因dhaT,并将该基因克隆到pMD19-TSimple载体。基因序列分析表明,dhaT基因全长为1158bp,与GenBank上已发表基因序列的同源性达99%,氨基酸同源性为100%。     

Implications of step-chilling on meat color investigated using proteome analysis of the sarcoplasmic protein fraction of beef longissimus lumborum muscle

In order to improve beef color and color stability,step-chilling(SC)was applied on excised bovine longissimus lumborum muscle,with chilling starting at 0–4℃ for 5 h,then holding the temperature at 12–18℃ for 6 h,followed by 0–4℃ again until 24 h post-mortem.pH and temperature were measured during rigor on SC loins as well as those subjected to routine chilling(RC,0–4℃,till 24 h post-mortem).Color L*,a*,b*values,metmyoglobin(MetM b)content,MetM b reducing ability(MRA)and NADH content were determined on samples aged for 1,7,and 14 d.Sarcoplasmic proteome analysis was only conducted on d 1 samples.The results showed muscles subjected to SC maintained a temperature at around 15℃ for 5 to 10 h post-mortem,and exhibited a slow temperature decline,but rapid pH decline.Beef steaks treated with SC had higher L*,a*,b* and chroma values than those of RC samples at 1 and 7 d chilled storage(0–4℃),while showing no significant difference for a*,b* and chroma values at d 14.The SC samples also exhibited a lower relative content of surface MetM b,higher MRA and NADH content,compared with RC beef steaks during storage,indicating the SC-treated beef showed an improved color stability.Eleven differential protein spots/nine proteins were identified by two-dimensional gel electrophoresis and mass spectrometry,and those proteins were mainly involved in redox,chaperone binding,metabolic and peroxidase activity.Oxidoreductases play a role in decreasing the oxidation-induced myoglobin oxidation and benefiting the production of NADH,and finally improving the colour of beef.Of these,pyruvate dehydrogenase E1 component subunit beta showed a positive correlation with color L*,a*,b*values and accounted for more than 60% of the variation in color values;this protein can be considered as a potential beef color biomarker.The present study provided valuable information for studies on the molecular mechanism of color improvement from step-chilling,as well as for identifying markers associated with beef color.     

芥菜不同类型品种及杂种的过氧化物酶同工酶酸性磷酸酶同工酶分析

利用聚丙烯酰胺凝胶电泳法研究了芥菜(Brassica iuncea L.)不同类型品种和杂种的过氧化物酶同工酶、酸性磷酸酶同工酶。结果表明:芥菜不同类型品种在两种同工酶上都表现出一定的差异,酸性磷酸酶同工酶还表现出一定规律性的差异。在杂合子中,过氧化物酶同工酶没有“杂种酶带”表现,而酸性磷酸酶同工酶有“杂种酶带”出现,且杂合子酶谱表现随双亲的酶谱变异而有所不同。正反交酶谱表现无差别。可以认为,这两种酶的编码基因都是核基因。酸性磷酸酶同工酶可作为“特征酶”在芥菜种质资源研究和育种实践中利用。