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Mitochondrial NAD(P)-dependent malic enzyme [EC 1.1.1.39, L-malate: NAD+ oxidoreductase (decarboxylating)] was purified from herring skeletal muscle to a specific activity of 8.2 mol/min/mg. The purification procedure involved chromatography on DEAE-cellulose, Red Agarose and a Sephacryl S-300 with a final recovery of 38% of enzyme activity. This enzyme catalyzes the oxidative decarboxylation of malate in the presence of either NAD or NADP in the presence of Mn2+. Some kinetic characteristics of this enzyme were determined. The pH optimum of activity is 7.0. ATP was shown to be a competitive inhibitor with malate. The inhibition by ATP displayed hyperbolic competitive kinetics with a Ki (ATP) of 0.28 mM in the presence of NAD and 0.75 mM in the presence of NADP. Fumarate reversed ATP inhibition.In vivo, regulation of NAD(P)-dependent malic enzyme might respond to changing levels of mitochondrial ATP and fumarate with the enzyme undergoing kinetic activation by an increase in the concentration of mitochondrial fumarate which could reverse enzyme inhibition by ATP. 相似文献
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Mitochondrial NAD(P)-dependent malic enzyme [EC 1.1.1.39, L-malate: NAD+ oxidoreductase (decarboxylating)] was purified from herring testicular tissue to a specific activity of 26.4 μmol NADH/min/mg
protein. Herring testicular tissue is one of the most abundant sources of this enzyme. The purification procedure involved
differential centrifugation of mitochondria and then chromatography on DEAE-Sephacel, Red Agarose and Sephacryl S-300. This
enzyme catalyzes the oxidative decarboxylation of malate in the presence of Mn2+ and either NAD or NADP. Under Vmax conditions the ratios for the rate of NAD/NADP reduction was 1.8. A study of the reductive carboxulation reaction indicated
that this enzyme reaction is reversible; at pH 7.0 the reverse reaction exhibited 22% of the activity of forward reaction.
Some kinetic characteristics of the enzyme were determined. ATP was found to be a competitive inhibitor with respect to malate.
Fumarate reversed ATP inhibition. Regulation of NAD(P)-dependent malic enzyme from herring testicular tissue mitochondria
could respond to changing levels of mitochondrial ATP and fumarate in vivo. 相似文献
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草莓果实采后NAD激酶活性与NAD（H）、NADP（H）含量及活性氧代谢的关系 总被引：6，自引：0，他引：6
【目的】研究草莓采后成熟衰老过程中NADK活性与NAD（H）、NADP（H）及活性氧代谢和膜氧化产物变化的关系，以探讨NADK在非跃变型果实成熟衰老过程中的作用，为调控果实的成熟衰老提供理论依据。【方法】将从果园采回的草莓果实贮藏于不同的温度下并进行每天取样，研究草莓果实在低温（4℃）、常温（20℃）贮藏期间成熟衰老过程中NAD激酶（NADK）活性及其底物NAD（H）、产物NADP（H）以及超氧阴离子O2- •过氧化氢（H2O2）、膜氧化产物丙二醛（MDA）含量的变化并分析NADK 与上述指标的关系。【结果】草莓果实在低温（4℃）贮藏时，其NADK活性比常温（20℃）贮藏的高，NAD（H）含量则相应比常温贮藏的低，NADP（H）含量则高于常温下的；同时，在常温贮藏期间果实O2- •生成速率和H2O2含量、膜氧化产物MDA含量均比低温贮藏的高，暗示NADK可能通过影响NAD（H）、NADP（H）的含量及比例来调控O2- •生成速率和H2O2含量，从而调控果实的成熟衰老。【结论】非跃变型果实草莓采后成熟衰老过程中，保持较高的NADK活性有利于延缓果实的成熟衰老，降低NADK活性可导致NAD和NAD（H）含量的积累，进而加速电子传递，产生大量的活性氧O2- •和H2O2，从而促进膜过氧化作用和积累较多的MDA，最终导致果实衰老变质。 相似文献
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香蕉果实贮藏冷害与NAD激酶活性变化的关系 总被引：2，自引：0，他引：2
以‘巴西'香蕉果实为试材,研究了果实采后在22℃和6℃下贮藏期间NAD激酶(NADK)活性与冷害发生情况,膜渗透性,丙二醛(MDA)含量,多酚氧化酶(PPO)以及苯丙氨酸解氨酶(PAL)活性变化的关系。结果表明,6℃下贮藏3d 时果实出现冷害症状, 11d时冷害加重,同时NAD激酶活性、膜渗透性、MDA含量和PPO活性均始终高于22℃贮藏的, 并在11d时达到高峰,而PAL活性则呈下降趋势,其活性始终低于22℃贮藏的。6℃下用NAD激酶激活剂氯化钙处理香蕉果实后,NADK受到激活,冷害程度加重,而用钙离子通道阻塞剂异博定（verapamil,Vp）处理后则抑制了NADK 活性,果实冷害症状延迟4 d 出现,冷害程度减弱,说明香蕉果实冷害发生与NADK 活性变化密切相关。 相似文献
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NAD(H)激酶是NADP(H)从头合成途径的关键物质,对生物体的生长、发育具有重要作用。本研究以稻瘟病菌中3个NAD(H)激酶蛋白为查询序列,搜索玉米大斑病菌蛋白质组序列库,结果发现在玉米大班病菌中均有同源序列,分别为127652、103052和163223。利用生物信息学的方法对3个基因的结构进行分析,结果表明只有163223为断裂基因,包含5个外显子和4个内含子,127652和103052为单外显子;163223和103052呈现酸性,而127652则呈现碱性;103052为稳定蛋白,而另两者为不稳定蛋白;通过高级结构分析发现这3种蛋白在N端一侧α螺旋较多,C端一侧延伸链和β转角较多,并且这3种蛋白质有2个在结构和功能上的相似的结构域。 相似文献
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《Journal of plant nutrition》2013,36(10-11):2187-2196
Abstract The present work aimed at evaluating in sunflower leaves: the relationship between Fe‐deficiency and the possible accumulation of H2O2; the activity of some extracellular enzymatic and non‐enzymatic systems involved in H2O2 production and scavenging. Iron‐deficient leaves exhibited a significant H2O2 accumulation, monitored at extracellular level by electron microscopy analysis. Such an increase in H2O2 level could derive from the significant decrease in extracellular ascorbate peroxidase (APX) activity, accompanied by a lower APX protein content detected by immuno‐electrophoresis. Also, extracellular ascorbic acid content was significantly affected by Fe‐deficiency, implying a reduced capacity for H2O2 scavenging. The contribute of peroxidases (PODs) involved in lignification process in keeping H2O2 levels under the toxic threshold was dependent on their subcellular location: the activity of covalently‐bound (CB) isoforms being increased while that of ionically‐bound (IB) ones decreased. NAD(P)H oxidation activity, known to produce H2O2, was found to decrease in apoplastic fluid and IB fraction and to slightly increase in the CB fraction. In conclusion, H2O2 accumulation observed in Fe‐deficient sunflower leaves seems to be mostly due to a reduced scavenging capacity of ASA, APX, and PODs, although the possibility of increased H2O2 production cannot be excluded. 相似文献
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叶片是植物进行光合作用的主要场所,其衰老由内源遗传发育信号和外界环境胁迫所启动,是一个非常复杂有序的调控过程。烟酰胺腺嘌呤二核苷酸(nicotinamide adenine dinucleotide,NAD)是脱氢酶的辅酶,在糖酵解、糖异生、三羧酸循环以及呼吸链等代谢中发挥着不可替代的作用。最新研究表明,水稻NAD生物合成参与调控沉默信息调控因子Sirtuins的生物活性、组蛋白H3K9去乙酰化、植物激素茉莉酸(JA)和叶片衰老。本文综述了有关水稻叶片衰老的细胞生理特征、Sirtuins酶活、NAD生物合成以及水稻早衰的OsSRT1-NAD调控途径和OsSRT1-Me OH-JA调控途径,以期阐明水稻叶片衰老的分子机理及其调控途径,为高产育种提供相应的理论参考。 相似文献
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Glutathione transferase (GST) was purified from the hindgut of grasshopper (Zonocerus variegatus) a polyphagous insect. The purified enzyme had a native molecular weight of 40 kDa and a subunit molecular weight of 19 kDa. The purified enzyme could conjugate glutathione (GSH) with 1-chloro-2,4-dinitrobenzene (CDNB), paranitrobenzylchloride, paranitrophenylacetate, 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBDCl), and 1,2-dichloro-4-nitrobenzene (DCNB) with specific activities of 3.3 ± 0.3, 0.49 ± 0.10, 0.10 ± 0.002, 1.2 ± 0.2, and 1.7 ± 0.4 μmol/min/mg protein, respectively. CDNB appears to be the best substrate with a specificity constant, kcat/Km, of 1.8 ± 0.1 × 10−4 M−1 S−1. The kinetic mechanism of Z. variegatus GST (zvGST) in the conjugation of GSH with some electrophilic substrates appears complex. Conjugation of GSH with DCNB was inhibited by high DCNB concentration, while with NBDCl, as the electrophilic substrates, different values of Km were obtained at high and low concentrations of the substrates. Cibacron blue, hematin, S-hexylglutathione, and oxidized glutathione inhibited the enzyme with I50 values of 0.057 ± 0.004, 0.80 ± 0.2, 33 ± 2 μM, and 5.2 ± 0.3 mM, respectively. The nature of inhibition by each of these inhibitors is either competitive or non-competitive at varying GSH or CDNB as substrates. NADH and NAD+ inhibited the enzyme with an I50 value of 0.4 ± 0.01 and 11 ± 1 mM, respectively. NADH at a concentration of 0.54 mM completely abolished the activity. As part of its adaptation, the flexible kinetic pathway of detoxication by zvGST may assist the organism in coping with various xenobiotics encountered in its preferred food plants. 相似文献
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Activities of NAD kinase(NADK)and NADP phosphatase and relationship between the two enzymes and temperature, respiration, ethylene production and trifluoperazine(TFP) were studied during ripening and senescence of strawberry and tomato frnits after harvest at 4℃and 20℃. The activity of NAD kinase in strawberry decreased slowly during first four days, then increased gradually. The NADP phosphatase activity increased at the second day, decreased the next day,then increased again. In tomato fruit, the activities of NAD kinase and NADP phosphatase increased at the second day, decreased with the ripening and senescence of the fruit. The change trend of NAD kinase and respiration in the two fruits were similar, the same were NADP phosphatase and ethylene production. TFP enhanced the activity of NAD kinase and had little effect on NADP phosphatase. Low temperature(4℃ ) activated the NAD kinase and reduced the activity of NADP phosphatase. These results indicated that the NAD kinase and NADP phosphatase were related to the ripening and senescence of strawberry and tomato fruits. The activation of NAD kinase probably postponed the ripening and senescence of the fruits. 相似文献