From Haemocuprein to Copper-Zinc Superoxide Dismutase: A History on the Fiftieth Anniversary of the Discovery of Haemocuprein and the Twentieth Anniversary of the Discovery of Superoxide Dismutase

Haemocuprein was discovered fifty years ago by T. Mann and D. Keilin as a copper protein of red blood cells, later named erythrocuprein. Superoxide dismutase was discovered twenty years ago by J.M. McCord and I. Fridovich as an enzymatic activity in preparations of carbonic anhydrase or myoglobin that inhibited the aerobic reduction of cytochrome c by xanthine oxidase. Astonishingly the superoxide dismutase proved to be haemocuprein. Around this time zinc was found in haemocuprein, in equimolar amount to the copper. Haemocuprein thus became copper-zinc superoxide dismutase after thirty years as an obscure cupropro-tein of red blood cells. This historical article is a tribute to the achievement of J.M. McCord and I. Fridovich. Their discovery of superoxide dismutase revolutionized the study of oxygen free-radicals in biochemistry.     

Comparative Analysis of Superoxide Dismutase Activity between Acute Pharmacological Models and a Transgenic Mouse Model of Huntington's Disease

We examined the activity of striatal superoxide dismutase (SOD) in two acute pharmacological models of Huntington's disease (HD), and compared it with SOD activity in the striata of mice transgenic for the HD mutation. Total SOD, and Cu/ZnSOD activities increased in young transgenic mice, but decreased in older (35 week) mice. We consider that the increased enzyme activity represents a compensatory mechanism to protect cells from free radical-induced damage, but the system becomes insufficient in older animals. Major decreases in SOD activity were also observed both after quinolinic acid and 3-nitropropionic acid intrastriatal injections. The present results indicate that in both types of HD models striatal oxidative damage occurs, and that it is associated with alterations in the cellular antioxidant system.     

超氧化物歧化酶在氧自由基所引起的癌变中的作用

超氧化物歧化酶是一类抗氧化酶,它能催化超氧阴离子自由基的歧化反应,对机体起保护作用。氧自由基在某些情况下会对机体产生损伤作用。文章就氧自由基的产生、氧自由基与致癌的关系,以及超氧化物歧化酶在氧自由基所引起的癌变中的作用等方面进行了综述。     

Effect of Silibinin on the Activity and Expression of Superoxide Dismutase in Lymphocytes from Patients with Chronic Alcoholic Liver Disease

The in vitro and in vivo effects of the naturally occuring Ravolignan hepatoprotective agent silibinin? on the expression and activity of superoxide dismutase (SOD) enzyme were studied in lymphocytes from patients with chronic alcoholic liver disease. In vitro incubation with silibinin in a concentration corresponding to the usual therapeutic dosage markedly increased the SOD — expression of lymphocytes as measured by Row-cytofluorimetry following staining with monoclonal anti-Cu, Zn-SOD — antibody and FITC-conjugated anti-mouse Ig. In vivo treatment with the drug restored the originally low SOD activity of the patients' lymphocytes. These data indirectly suggest that antioxidant activity might be one of the important factors in the hepatoprotective action of silibinin.     

活性氧所致超氧化物歧化酶肽链断裂的观察

探究活性氧所致铜锌超氧化物歧化酶（ＳＯＤ）肽链断裂的情况。将过氧化氢或抗坏血酸－Ｆｅ（Ⅲ）分别作用于马来酰亚胺标记的ＳＯＤ,然后用高效液相反相色谱（ＲＰＨＰＬＣ）分析,经１ｍｍｏｌ／ＬＨ２Ｏ２处理后ＳＯＤ用ＲＰ－ＨＰＬＣ分离出二个肽段,用顺磁共振检测显示只有一个肽段具有马来酰亚胺信号,经５ｍｍｏｌ／ＬＨ２Ｏ２处理后ＳＯＤ有四个肽段生成,其中有一个肽段具有马来酰亚胺信号,用５ｍｍｏｌ／Ｌ抗坏血酸和０．０１ｍｍｏｌ／ＬＦｅＣｌ３处理后ＳＯＤ有三个肽段生成,用５０ｍｍｏｌ／Ｌ抗坏血酸及１．０ｍｍｏｌ／ＬＦｅＣｌ３处理后ＳＯＤ也产生相同的三个肽段,只是肽段的量多．结果提示Ｈ２Ｏ２所致ＳＯＤ肽链断裂无“定点”现象,而抗坏血酸－Ｆｅ（Ⅲ）所致ＳＯＤ肽链断裂呈“定点”断裂。     

植物超氧化物歧化酶（SOD）的研究进展

超氧化物歧化酶(superoxide dismutase,SOD)在需氧原核生物和真核生物中广泛存在,是活性氧清除系统中第一个发挥作用的抗氧化酶。植物正常代谢过程和在各种环境胁迫下均能产生活性氧和自由基,活性氧和自由基的积累引起细胞结构和功能的破坏。SOD岐化超氧物阴离子自由基生成过氧化氢和分子氧,在保护细胞免受氧化损伤过程中具有十分重要的作用。本文综述了SOD的功能、在细胞中的分布、表达调控和与植物抗逆性的关系。 Abstract:Superoxide Dismutases (SODs) are ubiquitously expressed antioxidant enzyme in aerobic organisms and catalyze dismutation of superoxide anion to hydrogen and molecular oxygen,the first step in active oxygen-scavenging systems.SODs play a central role in protecting cells against the toxic effects of reactive oxygen species generated during normal cellular metabolic activity or as a result of various environmental stresses.This paper reviews the expression and regulation of Sod genes and their functional role(s) during development and in response to stresses.     

Bisphenol-A Can Inhibit the Enzymatic Activity of Human Superoxide Dismutase

Bisphenol-A (BPA), a synthetic xenoestrogen, is currently being used to produce a wide variety of consumer products. Humans as well as animals are exposed to this ubiquitous compound through ingestion, inhalation, and dermal exposure. The effect of this compound on superoxide dismutase (SOD), an antioxidant enzyme, isolated from human blood was studied using an enzyme inhibition assay. The mode of interaction of BPA on SOD was investigated using modeling and docking studies. Purified human SOD from erythrocytes was used to study the enzyme inhibition assay of BPA. Molecular level interactions of BPA on SOD were also analyzed by modeling and docking studies. Our study demonstrates that BPA has an inhibitory effect on SOD. The docking results showed that it could bind to the active site residues of SOD and could interfere with the catalytic activity of the enzyme. Our study reveals for the first time that BPA can directly inhibit the enzymatic activity of human SOD and thus impairs the free radical scavenging mechanism.     

Expression of Human Cu-Zn Superoxide Dismutase Gene in Transgenic Mice: Model for Gene Dosage Effect in Down Syndrome

It was suggested that increased Cu-Zn superoxide dismutase (SOD-I) might be involved in the various biological abnormalities found in Down's syndrome (DS) such as premature aging and Alzheimer-type neurological lesions. As a model system for testing this hypothesis we have developed two strains of transgenic mice carrying only one copy of the human SOD-I gene. In the first strain (TGI). no expression has been found by northern blot analysis. The second strain (TG2) exhibited human SOD-I mRNA and increased SOD-l activity in the brain (1.93 fold), in the heart (I.69 fold), thymus (I.49 fold) and to a lesser extent in muscle (1.25 fold), liver (1.19 fold), kidney (1.18 fold), spleen (1.35 fold), lung (1.26 fold) and erythrocytes (1.09 fold). In this strain, increased SOD-1 activity in the brain did not induce modifications in the seleno-dependent glutathione peroxidase. glutathione reductase and glutathione S-transferase activities. In brain homogenates, we have focused our studies on Tau proteins which are known to be the major antigenic components of paired helical filaments (P11F). both in DS and Alzheimer's disease. Our results suggested that, in our experimental conditions, the overexpression of SOD-I did not induce the modifications of Tau proteins similar to those seen during neurofibrillary degeneration.     

Decreased Catalase Activity but Unchanged Superoxide Dismutase Activity in Brains of Patients with Dementia of Alzheimer Type

Abstract: "Oxidative stress" may be of significance in the etiopathogenesis of dementia of Alzheimer type (DAT). Therefore, we measured activities of the enzymes superoxide dismutase (SOD) and catalase (CAT), which detoxicate reactive oxygen species. Enzyme activities were measured postmortem in basal ganglia, cortical, and limbic brain regions of patients with DAT and age-matched controls. SOD activity increased with age in basal nucleus of Meynert. However, there was no significant difference in SOD activity between DAT and controls. CAT activity was independent of age and postmortem time. There were significant reductions in CAT activity in parietotemporal cortex, basal ganglia, and amygdala in DAT compared with controls ( p < 0.05 to 0.01). Our findings are in line with the assumption that reactive oxygen species could contribute to the pathogenesis of DAT. Absence of these changes in basal nucleus of Meynert might reflect retrograde degeneration of cholinergic fibers.     

人锰超氧化物歧化酶cDNA的克隆、测序及表达

用逆转录-聚合酶链反应(RT-PCR)以人肝细胞总RNA为模板, 扩增了人锰超氧化物歧化酶(hMnSOD)的cDNA片段, 将此cDNA克隆到载体pGEM-T中.对重组质粒进行限制酶切分析和序列测定, 确定为含hMnSODcDNA的重组质粒将该hMnSODcDNA重组到表达载体pBV220内, 重组质粒在大肠杆菌DH5-α中表达hMnSOD, 表达产物占菌体总蛋白的14%, 具有持异性SOD酶活性.     

Expression of Human Cu-Zn Superoxide Dismutase Gene in Transgenic Mice: Model for Gene Dosage Effect in Down Syndrome

It was suggested that increased Cu-Zn superoxide dismutase (SOD-I) might be involved in the various biological abnormalities found in Down's syndrome (DS) such as premature aging and Alzheimer-type neurological lesions. As a model system for testing this hypothesis we have developed two strains of transgenic mice carrying only one copy of the human SOD-I gene. In the first strain (TGI). no expression has been found by northern blot analysis. The second strain (TG2) exhibited human SOD-I mRNA and increased SOD-l activity in the brain (1.93 fold), in the heart (I.69 fold), thymus (I.49 fold) and to a lesser extent in muscle (1.25 fold), liver (1.19 fold), kidney (1.18 fold), spleen (1.35 fold), lung (1.26 fold) and erythrocytes (1.09 fold). In this strain, increased SOD-1 activity in the brain did not induce modifications in the seleno-dependent glutathione peroxidase. glutathione reductase and glutathione S-transferase activities. In brain homogenates, we have focused our studies on Tau proteins which are known to be the major antigenic components of paired helical filaments (P11F). both in DS and Alzheimer's disease. Our results suggested that, in our experimental conditions, the overexpression of SOD-I did not induce the modifications of Tau proteins similar to those seen during neurofibrillary degeneration.     

CuZn-Superoxide Dismutase, Extracellular Superoxide Dismutase, and Glutathione Peroxidase in Blood from Individuals Homozygous for Asp90Ala CuZn-Superoxide Dismutase Mutation

Abstract: The Asp⁹⁰Ala CuZn-superoxide dismutase mutation is associated with amyotrophic lateral sclerosis (ALS) in both homo- and heterozygous form. We analyzed antioxidant enzymes in blood from 44 individuals homozygous and 114 individuals heterozygous for the Asp⁹⁰Ala mutation as well as 66 blood relatives carrying the wild-type allele only. Erythrocyte CuZn-superoxide dismutase activity was reduced by 9% in the homozygous individuals, confirming previous findings on a smaller cohort. The specific activity of Asp⁹⁰Ala mutant CuZn-superoxide dismutase in erythrocytes was equal to that of isolated mutant enzyme and slightly higher than that of isolated wild-type enzyme. There was no evidence for the presence of inactive mutant molecules in erythrocytes, and the lower activity is due to the occurrence of fewer active molecules. There were no significant differences between the groups in plasma extracellular superoxide dismutase content, and the erythrocyte glutathione peroxidase activities were virtually identical. Also, there were no differences in these parameters between homozygous individuals with or without ALS. There was no evidence for any association with ALS of a polymorphic extracellular superoxide dismutase mutation, Arg²¹³Gly. The absence of response of the blood antioxidant enzymes to the Asp⁹⁰Ala CuZn-superoxide dismutase mutation does not support the theory that the ALS-linked CuZn-superoxide dismutase mutations cause disease by increased oxidant stress.     

橄榄超氧化物歧化酶的分离纯化与性质研究

采用超声波破碎、硫酸铵分级沉淀、SephadexG-100和DE-52柱层析,从橄榄中分离纯化Cu,Zn-SOD,并对其部分性质进行分析鉴定。结果得到酶的比活力为906.3U/mg。该酶对H2O2和KCN敏感,而T氏液对酶活性没影响。紫外吸收峰在275nm处,PAGE蛋白和活性染色呈现3条相对应的谱带,相对分子质量约为31.63kD,亚基相对分子质量约为15.73kD。此酶对热稳定,在pH7~9范围内稳定。     

赤子爱胜蚓超氧化物歧化酶的纯化和部分性质研究

采用硫酸铵分级沉淀和柱层析的方法,从赤子爱胜蚓整体细胞抽提液内分离得到纯的铜锌超氧化物岐化酶（Ｃｕ,Ｚｎ－ＳＯＤ）。每１００ｇ蚯蚓得到的ＳＯＤ制品,总活力为１１１５０Ｕ,比活力为５１３８Ｕ／ｍｇ蛋白,回收率为２０％。铜锌超氧化物岐化酶呈淡蓝绿色,最大紫外吸收波长为２７０ｎｍ。测得该酶分子量为３３０００,亚基分子量为１６５００。该酶亚基由１５６个氨基酸残基组成,不含酪氨酸。     

重组人超氧化物歧化酶的基因克隆、表达及产物纯化研究

为了克隆人SOD-cDNA,构建表达载体,实现其在大肠杆菌中的高效稳定表达,通过抽提人肝组织总RNA,RT-PCR扩增人SOD cDNA,构建含rhSOD cDNA的表达质粒pLY-4/rhSOD,转化大肠杆菌JF1125进行表达研究。结果克隆到的rhSOD cDNA序列与文献报道一致,在宿主菌中获得高效表达,表达水平达68%以上;蛋白复性纯化工艺高效快速,rhSOD纯品纯度达98%以上,比活性达到2529u/mg;为用基因工程方法生产rhSOD打下基础。     

Catalase and Superoxide Dismutase in the Cells of Strictly Anaerobic Microorganisms

Strictly anaerobic microorganisms relating to various physiological groups were screened for catalase and superoxide dismutase (SOD) activity. All of the investigated anaerobes possessed SOD activity, necessary for protection against toxic products of oxygen reduction. High specific activities of SOD were found in Acetobacterium woodii and Acetobacterium wieringae. Most of the investigated clostridia and acetogens were catalase-negative. A significant activity of catalase was found in Thermohydrogenium kirishiense, in representatives of the genus Desulfotomaculum, and in several methanogens. Methanobrevibacter arboriphilus had an exceptionally high catalase activity after growth in medium supplemented with hemin. Hemin also produced a strong positive effect on the catalase activity in many other anaerobic microorganisms. In methanogens, the activities of the enzymes of antioxidant defense varied in wide ranges depending on the stage of growth and the energy source.     

Inhibition of Cell Growth by Overexpression of Manganese Superoxide Dismutase (MnSOD) in Human Pancreatic Carcinoma

Manganese superoxide dismutase (MnSOD) levels have been found to be low in human pancreatic cancer [Pancreas26, (2003), 23] and human pancreatic cancer cell lines [Cancer Res.63, (2003), 1297] when compared to normal human pancreas. We hypothesized that stable overexpression of pancreatic cancer cells with MnSOD cDNA would alter the malignant phenotype. MIA PaCa-2 cells were stably transfected with a pcDNA3 plasmid containing sense human MnSOD cDNA or containing no MnSOD insert by using the lipofectAMINE method. G418-resistant colonies were isolated, grown and maintained. Overexpression of MnSOD was confirmed in two selected clones with a 2-4-fold increase in MnSOD immunoreactive protein. Compared with the parental and neo control cells, the MnSOD-overexpressing clones had decreased growth rates, growth in soft agar and plating efficiency in vitro, while in vivo, the MnSOD-overexpressing clones had slower growth in nude mice. These results suggest that MnSOD may be a tumor suppressor gene in human pancreatic cancer.