大肠杆菌guaC基因的克隆及高效表达

以E.coli MC4100染色体DNA为模板使用PCR扩增技术克隆了鸟苷酸还原酶基因guaC，该基因全长1044bp，编码相对分子质量为37ku的蛋白质。将该基因直接克隆于PET—16b质粒的T7启动于下游，得到质粒pEXP1。转化E．coli BL21(DE3)，经异丙基硫代半乳糖苷(IPTG)诱导，获得了高效的表达。含表达质粒的菌体胞内粗提液经酶活分析表明，鸟苷酸还原酶的活性为不含质粒的宿主菌的80-85倍。     

人细胞色素P450氧化还原酶的克隆表达和抗体的制备

细胞色素P450还原酶(CPR)是人细胞色素P450酶系的电子供给者,对后者活性的发挥起到重要作用.本研究将从人胚胎cDNA文库中得到的人CPR的编码基因,连接入pT7450表达载体中,在大肠杆菌BL21(DE3)中高效表达并纯化.每升发酵液可得到纯化蛋白49 mg,占总蛋白含量的10%,以细胞色素C为底物检测酶活性,比活为65 u/mg.利用纯化的CPR作为抗原,常规免疫纯系大耳家兔,获得高效价、高特异性的抗血清,抗体滴度为1∶200 000(ELISA法),纯化后抗体应用于不同组织中CPR含量的评价,证明重组CPR及其抗体可用于细胞色素P450的体外药物代谢及细胞色素P450与CPR作用机理的研究.     

流动注射化学发光抑制法测定痕量亚硝酸根的研究

基于在碱性介质中,亚硝酸根对Lum inol-K3Fe(CN)6 体系发光反应具有强烈的抑制作用,建立了化学发光抑制快速测定痕量亚硝酸根的方法.测定NO2- 线性范围为1.0×10- 8 ～1.0×10- 6 g/m L,D.L= 5.6×10- 9 g/m L;对 4.0×10- 8g/m LNO-2 连续11 次测定的相对标准偏差为2.5% .     

Nitric oxide synthase reduces nitrite to NO under anoxia

Cultured bEND.3 endothelial cells show a marked increase in NO production when subjected to anoxia, even though the normal arginine pathway of NO formation is blocked due to absence of oxygen. The rate of anoxic NO production exceeds basal unstimulated NO synthesis in normoxic cells. The anoxic release of NO is mediated by endothelial nitric oxide synthase (eNOS), can be abolished by inhibitors of NOS and is accompanied by consumption of intracellular nitrite. The anoxic NO release is unaffected by the xanthine oxidase inhibitor oxypurinol. The phenomenon is attributed to anoxic reduction of intracellular nitrite by eNOS, and its magnitude and duration suggests that the nitrite reductase activity of eNOS is relevant for fast NO delivery in hypoxic vascular tissues. Received 20 August 2006; received after revision 21 September 2006; accepted 8 November 2006     

λλ双波长一阶导数紫外光度法测定环境水体中的亚硝酸盐氮

利用NO2^-在紫外区有特征吸收及｜dA／dλ｜与其浓度之间存在线性关系的特点，在232nm和214nm两波长处建立了一种用一元线性方程测定亚硝酸盐氮的新方法，并在环境水体的测定中取得令人满意的效果．     

溴酸钾-藏红T光度法测定痕量亚硝酸根

研究了在磷酸介质中 ,由于亚硝酸根催化作用 ,溴酸钾能够氧化藏红 T使之褪色 ,建立了测定痕量亚硝酸根的新方法 ,并探讨了这一催化反应的动力学条件 .其线性范围为 0～ 5.6 μg/ L,检出限为 2 .0× 1 0 - 8g/ L.用于纯净水中的亚硝酸根测定 ,结果满意     

All in the family: aldose reductase and closely related aldo-keto reductases

Aldose reductase catalyzes the first step in the polyol pathway and is thought to be involved in the pathogenesis of diabetic complications. In addition to polyol synthesis, aldose reductase may have multiple other activities that intersect with signal processing and oxidative defense mechanisms. Multiple aldose reductase-like proteins have been discovered to have structures and catalytic properties that broadly overlap those of aldose reductase. This chapter will summarize new insights into properties and functions of aldose reductase and closely related members of the aldo-keto reductase enzyme superfamily.     

Functional, biochemical and genetic diversity of prokaryotic nitrate reductases

Prokaryotic nitrate reduction can serve a number of physiological roles and can be catalysed by a number of biochemically distinct nitrate reductases. Three distinct nitrate reductase classes can be indentified in prokaryotes, NAS, NAR and NAP. NAS is located in the cytoplasmic compartment and participates in nitrogen assimilation. NAR is usually a three-subunit complex anchored to the cytoplasmic face of the membrane with its active site located in the cytoplasmic compartment and is involved in anaerobic nitrate respiration. NAP is a two-subunit complex, located in the periplasmic compartment, that is coupled to quinol oxidation via a membrane anchored tetraheme cytochrome. It shows considerable functional flexibility by participating in anaerobic respiration or redox energy dissipation depending on the organism in which it is found. The members of all three classes of enzymes bind the bis-molybdopterin guanine dinucleotide cofactor at the active site, but they differ markedly in the number and nature of cofactors used to transfer electrons to this site. Analysis of prokaryotic genome sequences available at the time of writing reveals that the different nitrate reductases are phylogenetically widespread.     

金属杂多酸聚吡咯修饰电极的制备及应用

在圆盘电极上 ,用电化学方法将单缺位 Dawson型金属取代磷钨杂多酸盐 [α2 - K8P2 W17O61(Ni( ) OH2 )· 17H2 O,α2 - K8P2 W17O61(Co( ) OH2 )· 16 H2 O]的阴离子 (P2 W17)掺杂到聚吡咯 (PPy)薄膜中 ,制成 PPy/ P2 W17/ GC化学修饰电极 .研究电极对 NO-2 电还原过程的催化机理 .其还原电流 ipc值与 NO-2 浓度在 7.4 1× 10 -6～ 8.2 4× 10 -5mol· L-1和 1.74× 10 -4～ 1.2 1× 10 -2 mol· L-1范围内 ,呈良好的线性关系 .其中检测下限为 3.10× 10 -7mol· L-1,可用于酸雨中 NO-2 的测定