MutaGEL Oxidative Stress II PCR Assay
MutaGEL OxStress II PCR Assay manufactured in Germany by Immundiagnostik
Size: 24 Samples
Method: PCR (RFLP)
Sample Type: DNA (e.g.whole blood, cheek swab)
Sample Size: 200 µL
For Research Use Only
Assay Background
Oxidative stress is defined as a pathological disequilibrium between the body’s molecules (e.g from food oxidation), which have free electrons and the reducing agents who normally react with them. The anions of superoxide and peroxide are mainly concerned and their surplus acts as aging cause and may especially generate diabetes or cancer. The protein superoxide dimutase 2 (manganese-dependent SOD2) catalyses radicals of superoxide (from many different metabolic sources) to peroxide; the protein catalase (CAT together with glutathion S-transferase) transforms peroxide to water and oxygen. The higher or lower serum concentration of reactive molecules – NO, ox-LDL, H2O2, and many others – as result of ecological agents (e.g., poor or well living conditions) is therefore “framed” by the a variation of their reducing enzymes. Firstly our MutaGEL OxStress II PCR Assay diagnoses the polymorphism Val16Ala of SOD2, which changes the enzymes recognition by mitochondria, in whom it acts on ROS (reactive oxygen species): diseased people with certain cancers have more often valine alleles of SOD2 than healthy people (the relative abundance of Val/Val homozygotes in lung cancer patients is 1,7x the number in healthy persons; in bladder cancer the deviation is still higher). Because valine alleles are 1/3 less active than alanine alleles, its reduced decomposition of oxidative stress molecules can generate cancer. The results are complicated by the fact, that the alanine allele of SOD2 may also act as arepathogenic factor in breast cancer (in unfavourable circumstances like with women with high body mass index and with many menstruation years), and this may point on a neoplastic potential of ROS metabolites produced by the more active enzyme.
Secondly, we diagnose the promotor polymorphism C-262T of human catalase – the enzyme acts downstream of SOD2 (s.a.) -, which changes the protein’s concentration in erythrocytes and blood: each T allele of a person increases the activity. This is generating a protective effect of its counterpart with a C against breast or pancreas cancer and here also a mirror effect exists with a pathogenic effect for CC homozygotes in the pathogenesis of diabetic neuropathy. Thus the enzymes allelic variants in oxidative stress generate pathogenic or protective effects depending on the examined disease. A further influence on the reduction of oxidative stress comes from the null mutants of glutathione S-transferase M1 and T1 (see PCR kit Mutagel GST M1/T1) and sequence changes in the genes for endothelial NO synthase + NAD(P)H oxidase (see PCR kit Mutagel Oxstress I), (M.E.).
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决定酶促反应最大速度Vm的因素是什么？是同一底物，尽管加不同种酶，其Vmax都相同吗?可以画曲线图解释一下吗？可是根据米曼氏方程来看又有点矛盾，Km变大，Vmax也需变大，才可使v变大。所以题目所述到底应该怎么理解呢
调节至最适合温度（一般是37度），合适的酸碱度时，酶的活性最高
我是新手，来到丁香园看到有这么多的热心人，感到很高兴！！我也有很多问题需要大家的帮助！！实验已经作了一年了可是毫无进展，心里很是着急！！
我的课题是以外消旋的苯基乙二醇为底物，用假丝酵母催化生成手性纯的S-型苯基乙二醇，由于是老课题，所以目标是提高转化反应的底物浓度和菌体的使用批次（目前菌体使用一批后便不能再使用）。
我曾试过很多种方法，但均效果不大！我试着在转化过程中添加醛类，酮类，醇类作为辅助底物，增加菌体的使用批次。可效果不好，特别是添加了醇类后还有的起了反作用，因为我的这个转化过程中涉及到NAD和NADPH的再生。（其转化过程是酵母先催化将外消旋的苯基乙二醇变为酮，再将酮还原为醇，经过这一过程就将外消旋的苯基乙二醇变为手性纯的S-型了）
我还试过用固定化的方法，海藻酸钙包埋法，可是这样底物浓度就更低了！！
我现在不知道下一步该如何做了，很着急，请大家帮帮忙。谢谢了！！谢谢
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