小麦幼苗中过氧化物歧化酶活性与幼苗脱水忍耐力相关性的研究

本文着重探讨当前国际上提出的一种植物抗旱机制的新理论——即生物自由基与植物保护酶系统间相互作用的理论。在水分逆境情况下,植物细胞内自由基产生和清除的平衡遭到破坏,过剩的自由基会伤害细胞膜系统。过氢化物歧化酶是氧自由基的清除剂,因此它的活性的高低应与植物的抗旱能力密切相关。以小麦幼苗为材料,采用光化学方法测定了过氧化物歧化酶活性,同时测定幼苗的脱水耐受力,发现在不同组织和器官中,在不同发育阶段的幼苗中,二者均存在有正相关性。此外,还测定了幼苗脱水前后过氧化物歧化酶活性的变化,发现酶活性在脱水处理时升高。我们推测小麦幼苗经受干旱的过程中,细胞内可能包含生物自由基的有害作用。     

过氧化氢对铜锌超氧化物歧化酶结构损伤作用的进一步研究

我们曾观察到H_2O_2可使牦牛红细胞超氧化物歧化酶的活性下降及某些理化性质发生变化。在此基础上,进一步确证H_2O_2可使该酶结构受到损伤,其依据为H_2O_2作用后,酶的圆二色谱变化显示其β-折叠含量减少,无规则卷曲含量增加;酶分子中有二酪氨酸生成;组氨酸、精氨酸、赖氨酸、丝氨酸和苏氨酸含量相对减少,而天冬氨酸、谷氨酸和甘氨酸含量相对增加;用2.4-二硝基苯肼试剂可测得羰基生成;SDS-PAGE测出酶的部分亚基被降解为小肽段;用荧光胺法测得自由氨基量的增加。本文对上述变化的机理进行了探讨。     

Cu,Zn Superoxide Dismutase Maturation and Activity Are Regulated by COMMD1

The maturation and activation of the anti-oxidant Cu,Zn superoxide dismutase (SOD1) are highly regulated processes that require several post-translational modifications. The maturation of SOD1 is initiated by incorporation of zinc and copper ions followed by disulfide oxidation leading to the formation of enzymatically active homodimers. Our present data indicate that homodimer formation is a regulated final step in SOD1 maturation and implicate the recently characterized copper homeostasis protein COMMD1 in this process. COMMD1 interacts with SOD1, and this interaction requires CCS-mediated copper incorporation into SOD1. COMMD1 does not regulate disulfide oxidation of SOD1 but reduces the level of SOD1 homodimers. RNAi-mediated knockdown of COMMD1 expression results in a significant induction of SOD1 activity and a consequent decrease in superoxide anion concentrations, whereas overexpression of COMMD1 exerts exactly the opposite effects. Here, we identify COMMD1 as a novel protein regulating SOD1 activation and associate COMMD1 function with the production of free radicals.     

过氧化氢对铜锌超氧化物歧化酶活性及某些理化性质的影响

用H_2O_2作用于牦牛红细胞铜锌超氧化物歧化酶。观察到酶活性随H_2O_2浓度升高及作用时间增加而下降;酶分子连接的铜和锌有所丢失;PAGE图谱中三条酶活性带成为四条酶活性带;等电点下降;680nm处表征二价铜光学性质的可见光吸收减弱;紫外吸收增加,表现为增色效应;内源性荧光减弱;在含有3.0mol/LKCl的PH3.8—5.4琥珀酸缓冲液中溶解度下降;酶对胰蛋白酶水解的敏感性增加。     

Histochemical Localization of Superoxide Dismutase Activity in Rat Brain

Histochemical localization of superoxide anion (O₂^·−) scavenging activity in rat brain was visualized by the tissue-blotting technique. The activity was thought to mainly depend on Cu/Zn-SOD, because the localization of the activity was identical with the immunohistochemistry of Cu/Zn-SOD and the localization of its mRNA in the brain. Moreover, the activity was dramatically decreased after treatment of Cu (I) chelater. The activity was detected in pyramidal cells of the cortex, granular, and mitral cells of the olfactory bulbs, pyramidal cell layer CA1 to CA3, and dentate gyrus of hippocampus formation and granular cells of the cerebellum. Moreover, the activity was detected in the pontine nuclei of brain stem. Olfactory bulbs, hippocampus, and cerebellum were believed to be bestowed high brain functions, i.e., long-term potentiation and long-term depression. A part of the function was regulated by a retrograde neurotransmitter, nitric oxide (^·NO). Our findings suggest that the SOD is colocalized with NO synthase in olfactory bulbs, hippocampus, and cerebellum, where ^·NO plays the important roles. In contrast, low SOD activity was observed in the axonal neurofiber bundles, although the regions contain a lot of membrane lipids, which was thought to be peroxidized by O₂^·− and related radicals such as ^·OH in the regions. From these findings, it was suggested that the SOD did not only play a role in protecting the neurons from endogenously formed O₂^·−, but also play a role in preservation of beneficial natures of ^·NO in the brain.     

Change of Cu,Zn-superoxide Dismutase Activity of Guinea Pig Lung in Experimental Asthma

Correlation between the level of reactive oxygen species (ROS) generated by airway inflammatory cells and superoxide dismutase (SOD) activity of pulmonary tissue during an asthma attach was investigated in a guinea pig model of allergic asthma. In addition, the influence of SOD inhibition by diethyldithiocarbamate (DDC, Cu-chelating agent) on the airway was investigated in terms of pulmonary function during an asthma attach. Relative to controls, the capacity of bronchoalveolar lavage fluid (BAL) cells to release ROS was significantly increased in guinea pigs sensitized with ovalbumin (OA) as the antigen, and significantly increased in guinea pigs with an asthma attack provoked by the inhalation of OA. SOD activity was increased significantly in the antigen-sensitized group. The asthma provocation group showed a tendency for increase in total SOD activity, compared with the sensitization group, whose increase was dependent on the increase in copper, zinc-SOD (Cu, Zn-SOD) activity. Pretreatment with DDC increased the severity and duration of the asthma attack. These results were indicated that Cu, Zn-SOD was closely involved in the asthma process, particularly in the scavenging of oxygen radicals secreted from BAL cells.     

Superoxide Dismutase Concentration and Activity in Familial Amyotrophic Lateral Sclerosis

Abstract: Some cases of autosomal-dominant familial amyotrophic lateral sclerosis (FALS) have been associated with mutations in SOD1 , the gene that encodes Cu/Zn superoxide dismutase (Cu/Zn SOD). We determined the concentrations (µg of Cu/Zn SOD/mg of total protein), specific activities (U/µg of total protein), and apparent turnover numbers (U/µmol of Cu/Zn SOD) of Cu/Zn SOD in erythrocyte lysates from patients with known SOD1 mutations. We also measured the concentrations and activities of Cu/Zn SOD in FALS patients with no identifiable SOD1 mutations, sporadic ALS (SALS) patients, and patients with other neurologic disorders. The concentration and specific activity of Cu/Zn SOD were decreased in all patients with SOD1 mutations, with mean reductions of 51 and 46%, respectively, relative to controls. In contrast, the apparent turnover number of the enzyme was not altered in these patients. For the six mutations studied, there was no correlation between enzyme concentration or specific activity and disease severity, expressed as either duration of disease or age of onset. No significant alterations in the concentration, specific activity, or apparent turnover number of Cu/Zn SOD were detected in the FALS patients with no identifiable SOD1 mutations, SALS patients, or patients with other neurologic disorders. That Cu/Zn SOD concentration and specific activity are equivalently reduced in erythrocytes from patients with SOD1 mutations suggests that mutant Cu/Zn SOD is unstable in these cells. That concentration and specific activity do not correlate with disease severity suggests that an altered, novel function of the enzyme, rather than reduction of its dismutase activity, may be responsible for the pathogenesis of FALS.     

银杏叶愈伤组织和叶中超氧化物歧化酶、总黄酮类抗氧化活性剂活性的比较分析

以银杏（Ginkgo biloba L.）叶片为外植体进行愈伤组织的诱导和培养,筛选到适合银杏叶片愈伤组织诱导和继代的培养基。以此愈伤组织为研究材料,比较了叶片愈伤组织和叶片中两种活性成分超氧化物歧化酶（SOD）、黄酮的活性差异。结果表明,银杏叶片愈伤组织中SOD的活性明显比叶片中的SOD活性高,同时愈伤组织中SOD同工酶谱带也比叶中有更多表达,但愈伤组织中总黄酮含量却比叶片中的含量低。实验证实SOD和黄酮类物质具有清除超氧阴离子的作用,表明银杏叶片愈伤组织和叶片中具有不同的占主导作用的抗氧化活性的机制。     

Insights into the Role of the Unusual Disulfide Bond in Copper-Zinc Superoxide Dismutase

The functional and structural significance of the intrasubunit disulfide bond in copper-zinc superoxide dismutase (SOD1) was studied by characterizing mutant forms of human SOD1 (hSOD) and yeast SOD1 lacking the disulfide bond. We determined x-ray crystal structures of metal-bound and metal-deficient hC57S SOD1. C57S hSOD1 isolated from yeast contained four zinc ions per protein dimer and was structurally very similar to wild type. The addition of copper to this four-zinc protein gave properly reconstituted 2Cu,2Zn C57S hSOD, and its spectroscopic properties indicated that the coordination geometry of the copper was remarkably similar to that of holo wild type hSOD1. In contrast, the addition of copper and zinc ions to apo C57S human SOD1 failed to give proper reconstitution. Using pulse radiolysis, we determined SOD activities of yeast and human SOD1s lacking disulfide bonds and found that they were enzymatically active at ∼10% of the wild type rate. These results are contrary to earlier reports that the intrasubunit disulfide bonds in SOD1 are essential for SOD activity. Kinetic studies revealed further that the yeast mutant SOD1 had less ionic attraction for superoxide, possibly explaining the lower rates. Saccharomyces cerevisiae cells lacking the sod1 gene do not grow aerobically in the absence of lysine, but expression of C57S SOD1 increased growth to 30–50% of the growth of cells expressing wild type SOD1, supporting that C57S SOD1 retained a significant amount of activity.     

煤尘对小鼠肺组织铜蓝蛋白和SOD活性的影响

目的通过接触煤尘后小鼠肺组织中铜蓝蛋白和SOD活性的改变情况,探讨铜蓝蛋白和SOD活性在煤尘肺发展中的意义。方法将50只小鼠随机分成空白对照组、实验对照组和1 h2、h3、h剂量组,采用动式呼吸道染毒,连续染毒12 d,染毒结束后将小鼠处死,测定其肺组织铜蓝蛋白和SOD的活性。结果小鼠脏器系数各组之间无差异;与对照组相比,各剂量组小鼠肺组织SOD活性均降低,差异显著（P〈0.05）,且空白对照与实验对照无差异（P〈0.05）;铜蓝蛋白各组之间无差异。结论短期接触一定浓度煤尘会引起SOD活性的改变。     

The Clinical and Laboratory Studies of Superoxide Dismutase Activity in the Human Whole Blood with Early Gastric Cancer

In this study, we tried to evaluate the usefulness of Superoxide Dismutase (SOD) activity in detecting gastric cancer. SOD was determined according to M. Minami and H. Yoshikawa. a simple and inexpensive assay method. For II fibrognstroscopy and pathological proved gastric cancer cases, the average levels of SOD activity was found significantly lower than 30 patients with gastric diseases, 8 patients with atypical hyperplasia and 32 controls.     

Rapid Communication: Cu/Zn Superoxide Dismutase Activity in Familial and Sporadic Amyotrophic Lateral Sclerosis

Abstract: Amyotrophic lateral sclerosis (ALS) is a degenerative motor neuron disease that is inherited as an autosomal dominant trait in ~ 10% of cases. Recently we and others identified several single-base mutations in the Cu/Zn superoxide dismutase (SOD1) gene in patients with familial ALS (FALS). Using single-strand conformational polymorphism, we studied the C to G mutation in exon 2 of the SOD1 gene (resulting in a leucine to valine substitution in position 38) in affected and unaffected members of a large Belgian family with FALS. We measured the SOD1 activity in red blood cell lysates in 14 members of this family, including the only surviving clinically affected patient. SOD1 activity of the family members carrying the mutation was less than half that of members without the mutation. In addition, in 11 patients with sporadic ALS and 11 age- and sex-matched controls, red blood cell SOD1 activity was normal. These studies indicate that SOD1 activity is reduced in these FALS patients but not in sporadic ALS patients. Moreover, this SOD1 enzyme abnormality is detectable years before onset of clinical ALS in carriers of this FALS mutation.     

Changes in Superoxide Dismutase Activity in Liver and Lung of Old Rats

Superoxide dismutase activity was measured in liver and lung from 3 and 24 month-old rats. Both total SOD and Mn-SOD activity decreased significantly in the liver of old rats. Recent results from our laboratory have indicated that during aging, the activity of Cu/Zn-SOD decreases in rat liver and that there is an accumulation of altered protein. It was also shown that the old Cu/Zn-SOD had one histidine fewer than the young one. In the present study, the immunoprecipitation experiments showed that the amount of immunoprecipitable Mn-SOD from liver of old rats was greater than from young ones, but when amino acid residues were measured in purified young and old Mn-SOD from liver, no change was observed. In lung, no significant age-related differences in total SOD, Cu/Zn-SOD and Mn-SOD activity were found, nor was there accumulation of altered protein during aging.     

Changes in Superoxide Dismutase Activity in Liver and Lung of Old Rats

Superoxide dismutase activity was measured in liver and lung from 3 and 24 month-old rats. Both total SOD and Mn-SOD activity decreased significantly in the liver of old rats. Recent results from our laboratory have indicated that during aging, the activity of Cu/Zn-SOD decreases in rat liver and that there is an accumulation of altered protein. It was also shown that the old Cu/Zn-SOD had one histidine fewer than the young one. In the present study, the immunoprecipitation experiments showed that the amount of immunoprecipitable Mn-SOD from liver of old rats was greater than from young ones, but when amino acid residues were measured in purified young and old Mn-SOD from liver, no change was observed. In lung, no significant age-related differences in total SOD, Cu/Zn-SOD and Mn-SOD activity were found, nor was there accumulation of altered protein during aging.     

Superoxide Dismutase Activity, Oxidative Damage, and Mitochondrial Energy Metabolism in Familial and Sporadic Amyotrophic Lateral Sclerosis

The cause of neuronal death in amyotrophic lateral sclerosis (ALS) is unknown. Recently, it was found that some patients with autosomal-dominant familial ALS (FALS) have point mutations in the gene that encodes Cu/Zn superoxide dismutase (SOD1). In this study of postmortem brain tissue, we examined SOD activity and quantified protein carbonyl groups, a marker of oxidative damage, in samples of frontal cortex (Brodmann area 6) from 10 control patients, three FALS patients with known SOD1 mutations (FALS-1), one autosomal-dominant FALS patient with no identifiable SOD1 mutations (FALS-0), and 11 sporadic ALS (SALS) patients. Also, we determined the activities of components of the electron transport chain (complexes I, II-III, and IV) in these samples. The cytosolic SOD activity, which is primarily SOD1 activity, was reduced by 38.8% (p < 0.05) in the FALS-1 patients and not significantly altered in the SALS patients or the FALS-0 patient relative to the control patients. The mitochondrial SOD activity, which is primarily SOD2 activity, was not significantly altered in the FALS-1, FALS-0, or SALS patients. The protein carbonyl content was elevated by 84.8% (p < 0.01) in the SALS patients relative to the control patients. Finally, the complex I activity was increased by 55.3% (p < 0.001) in the FALS-1 patients relative to the control patients. These results from cortical tissue demonstrate that SOD1 activity is reduced and complex I activity is increased in FALS-1 patients and that oxidative damage to proteins is increased in SALS patients.     

Oxidation of the Tryptophan 32 Residue of Human Superoxide Dismutase 1 Caused by Its Bicarbonate-dependent Peroxidase Activity Triggers the Non-amyloid Aggregation of the Enzyme

The role of oxidative post-translational modifications of human superoxide dismutase 1 (hSOD1) in the amyotrophic lateral sclerosis (ALS) pathology is an attractive hypothesis to explore based on several lines of evidence. Among them, the remarkable stability of hSOD1^WT and several of its ALS-associated mutants suggests that hSOD1 oxidation may precede its conversion to the unfolded and aggregated forms found in ALS patients. The bicarbonate-dependent peroxidase activity of hSOD1 causes oxidation of its own solvent-exposed Trp³² residue. The resulting products are apparently different from those produced in the absence of bicarbonate and are most likely specific for simian SOD1s, which contain the Trp³² residue. The aims of this work were to examine whether the bicarbonate-dependent peroxidase activity of hSOD1 (hSOD1^WT and hSOD1^G93A mutant) triggers aggregation of the enzyme and to comprehend the role of the Trp³² residue in the process. The results showed that Trp³² residues of both enzymes are oxidized to a similar extent to hSOD1-derived tryptophanyl radicals. These radicals decayed to hSOD1-N-formylkynurenine and hSOD1-kynurenine or to a hSOD1 covalent dimer cross-linked by a ditryptophan bond, causing hSOD1 unfolding, oligomerization, and non-amyloid aggregation. The latter process was inhibited by tempol, which recombines with the hSOD1-derived tryptophanyl radical, and did not occur in the absence of bicarbonate or with enzymes that lack the Trp³² residue (bovine SOD1 and hSOD1^W32F mutant). The results support a role for the oxidation products of the hSOD1-Trp³² residue, particularly the covalent dimer, in triggering the non-amyloid aggregation of hSOD1.     

凤眼莲超氧物歧化酶活性与抗寒性的关系

低温使凤眼莲叶内SOD活性降低,同时叶组织电解质外渗增加和TTG的还原力降低。用适量的联吡啶季胺盐(Paraquat)处理凤眼莲叶片,使超氧物歧化酶活性增强,则能减轻低温引起的伤害。试验提供了超氧物歧化酶与抗寒性有关的直接证据.     

Superoxide Triggers an Acid Burst in Saccharomyces cerevisiae to Condition the Environment of Glucose-starved Cells

Although yeast cells grown in abundant glucose tend to acidify their extracellular environment, they raise the pH of the environment when starved for glucose or when grown strictly with non-fermentable carbon sources. Following prolonged periods in this alkaline phase, Saccharomyces cerevisiae cells will switch to producing acid. The mechanisms and rationale for this “acid burst” were unknown. Herein we provide strong evidence for the role of mitochondrial superoxide in initiating the acid burst. Yeast mutants lacking the mitochondrial matrix superoxide dismutase (SOD2) enzyme, but not the cytosolic Cu,Zn-SOD1 enzyme, exhibited marked acceleration in production of acid on non-fermentable carbon sources. Acid production is also dramatically enhanced by the superoxide-producing agent, paraquat. Conversely, the acid burst is eliminated by boosting cellular levels of Mn-antioxidant mimics of SOD. We demonstrate that the acid burst is dependent on the mitochondrial aldehyde dehydrogenase Ald4p. Our data are consistent with a model in which mitochondrial superoxide damage to Fe-S enzymes in the tricarboxylic acid (TCA) cycle leads to acetate buildup by Ald4p. The resultant expulsion of acetate into the extracellular environment can provide a new carbon source to glucose-starved cells and enhance growth of yeast. By triggering production of organic acids, mitochondrial superoxide has the potential to promote cell population growth under nutrient depravation stress.     

A Circadian Rhythm in the Activity of Superoxide Dismutase in the Photosynthetic Alga Gonyaulax polyedra

The activity of superoxide dismutase in cell-free extracts of Gonyaulax made at different times of day and night was found to be three to four times higher during the day. This rhythm continued in cells kept in constant light, indicating that the regulation can be attributed to the cellular circadian clock.