Superoxide Dismutase and Oxygen Toxicity in a Eukaryote

Saccharomyces cerevisiae var. ellipsoideus contained 6.5 times more superoxide dismutase and 2.3 times more catalase when grown under 100% O(2) than when grown anaerobically. Growth under oxygen caused equal increases in both the cyanide-sensitive and the cyanide-insensitive superoxide dismutases of this organism. Experience with other eukaryotes has shown that cyanide sensitivity is a property of the cupro-zinc superoxide dismutase of the cytosol, whereas cyanide insensitivity is a property of the corresponding mangani-enzyme found in mitochondria. Cu(2+), which has been shown to increase the radioresistance of yeast, also caused an increase of both of the superoxide dismutases of S. cerevisiae. Yeast which had been grown under 1 atm of O(2) were more resistant toward the lethal effects of 20 atm of O(2) than were yeast which had been grown in the absence of O(2). Escherichia coli K-12 his(-) responded to growth under 1 atm of O(2) by increasing its content of catalase and of peroxidase, but not of superoxide dismutase. This contrasts with E. coli B, which was previously shown to respond to O(2) by a striking increase in superoxide dismutase. E. coli K-12 his(-) did not gain resistance toward 20 atm of O(2) because of having been grown under 1 atm of O(2). Once again, this contrasts with the behavior of E. coli B. These data indicate that, in both prokaryotes and in eukaryotes, superoxide dismutase is an important component of the defenses against oxygen toxicity.     

Induction of Superoxide Dismutase by Molecular Oxygen

Oxygen induces superoxide dismutase in Streptococcus faecalis and in Escherichia coli B. S. faecalis grown under 20 atm of O(2) had 16 times more of this enzyme than did anaerobically grown cells. In the case of E. coli, changing the conditions of growth from anaerobic to 5 atm of O(2) caused a 25-fold increase in the level of superoxide dismutase. Induction of this enzyme was a response to O(2) rather than to pressure, since 20 atm of N(2) was without effect. Induction of superoxide dismutase was a rapid process, and half of the maximal level was reached within 90 min after N(2)-grown cells of S. faecalis were exposed to 20 atm of O(2) at 37 C. S. faecalis did not contain perceptible levels of catalase under any of the growth conditions investigated by Stanier, Doudoroff, and Adelberg (23), and the concentration of catalase in E. coli was not affected by the presence of O(2) during growth. S. faecalis, which had been grown under 100% O(2) and which therefore contained an elevated level of superoxide dismutase, was more resistant of 46 atm of O(2) than were cells which had been grown under N(2). E. coli grown under N(2) contained as much superoxide dismutase as did S. faecalis grown under 1 atm of O(2). The E. coli which had been grown under N(2) was as resistant to the deleterious effects of 50 atm of O(2) as was S. faecalis which had been grown under 1 atm of O(2). These results are consistent with the proposal that the peroxide radical is an important agent of the toxicity of oxygen and that superoxide dismutase may be a component of the systems which have been evolved to deal with this potential toxicity.     

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。     

Cu,Zn-Superoxide Dismutase Increases Toxicity of Mutant and Zinc-deficient Superoxide Dismutase by Enhancing Protein Stability

When replete with zinc and copper, amyotrophic lateral sclerosis (ALS)-associated mutant SOD proteins can protect motor neurons in culture from trophic factor deprivation as efficiently as wild-type SOD. However, the removal of zinc from either mutant or wild-type SOD results in apoptosis of motor neurons through a copper- and peroxynitrite-dependent mechanism. It has also been shown that motor neurons isolated from transgenic mice expressing mutant SODs survive well in culture but undergo apoptosis when exposed to nitric oxide via a Fas-dependent mechanism. We combined these two parallel approaches for understanding SOD toxicity in ALS and found that zinc-deficient SOD-induced motor neuron death required Fas activation, whereas the nitric oxide-dependent death of G93A SOD-expressing motor neurons required copper and involved peroxynitrite formation. Surprisingly, motor neuron death doubled when Cu,Zn-SOD protein was either delivered intracellularly to G93A SOD-expressing motor neurons or co-delivered with zinc-deficient SOD to nontransgenic motor neurons. These results could be rationalized by biophysical data showing that heterodimer formation of Cu,Zn-SOD with zinc-deficient SOD prevented the monomerization and subsequent aggregation of zinc-deficient SOD under thiol-reducing conditions. ALS mutant SOD was also stabilized by mutating cysteine 111 to serine, which greatly increased the toxicity of zinc-deficient SOD. Thus, stabilization of ALS mutant SOD by two different approaches augmented its toxicity to motor neurons. Taken together, these results are consistent with copper-containing zinc-deficient SOD being the elusive “partially unfolded intermediate” responsible for the toxic gain of function conferred by ALS mutant SOD.     

极谱氧电极法测定超氧化物歧化酶活性的改进

对SOD的极谱氧电极测定法做了如下修改:a.室温测定,b.酶活性用标准SOD标定,c.反应在磷酸缓冲液中进行,d.增大邻苯三酚的用量.改进后克服了易在电极薄膜表面产生气泡等问题,测定灵敏度及线性范围增大.     

活性氧所致超氧化物歧化酶疏水性的变化

用抗坏血酸-Fe(Ⅲ)和过氧化氢分别作用于铜锌超氧化物歧化酶(SOD),经疏水层析分离得到亲水型和疏水型SOD.用胰蛋白酶和胃蛋白酶分别作用于天然SOD,亲水型SOD及疏水型SOD,结果表明疏水型SOD较亲水型SOD及天然SOD易被降解,提示活性氧氧化修饰后的SOD对蛋白水解酶敏感性提高与其疏水性增高有关.     

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。     

Superoxide Dismutase and Oxygen Metabolism in Streptococcus faecalis and Comparisons with Other Organisms

Streptococcus faecalis contains a single superoxide dismutase that has been purified to homogeneity with a 55% yield. This enzyme has a molecular weight of 45,000 and is composed of two subunits of equal size. It contains 1.3 atoms of manganese per molecule. Its amino acid composition was determined and is compared with that for the superoxide dismutases from Escherichia coli, Streptococcus mutans, and Mycobacterium lepraemurium. When used as an antigen in rabbits, the S. faecalis enzyme elicited the formation of a precipitating and inhibiting antibody. This antibody cross-reacted with the superoxide dismutase present in another strain of S. faecalis, but neither inhibited nor precipitated the superoxide dismutases in a wide range of other bacteria, including several other streptococci, such as S. pyogenes, S. pneumoniae, and S. lactis. The inhibiting antibody was used to suppress the superoxide dismutase activity present in cell extracts of S. faecalis and thus allow the demonstration that 17% of the total oxygen consumption by such extracts, in the presence of reduced nicotinamide adenine dinucleotide, was associated with the production of O(2) (-). A variety of bacterial species were surveyed for their content of superoxide dismutases. The iron-containing enzyme was distinguished from the manganese-containing enzyme through the use of H(2)O(2), which inactivates the former more readily than the latter. Some of the bacteria appeared to contain only the iron enzyme, others only the manganese enzyme, and still others both. Indeed, some had multiple, electrophoretically distinct superoxide dismutases in both categories. There was no discernible absolute relationship between the types of superoxide dismutases in a particular organism and their Gram-stain reaction.     

Evidence for Superoxide Dismutase and Catalase in Mollicutes and Release of Reactive Oxygen Species

The presence of superoxide dismutase was demonstrated in 21 strains of mollicutes, including achuloplas-mas, mycoplasmas and ureaplasmas. Additionally, catalase activities were demonstrated in nearly 50% of the cell lysates. whereas no peroxide activities were detectable. The production of O₂-and H₂O₂ with glucose as substrate was demonstrated for 8 strains of 10 strains tested. Anaerobic mycoplasmas showed the highest amount of radical production, whereas superoxide dismutase and catalase activities were in the range of activities estimated for aerobic mollicutes. Some pathogenic strains additionally released compounds into the culture medium, which stimulated O₂-production by PMNs.     

Superoxide Dismutase in Plants

Superoxide dismutases (SODs) are metal-containing enzymes that catalyze the dismutation of superoxide radicals to oxygen and hydrogen peroxide. The enzyme has been found in all aerobic organisms examined where it plays a major role in the defense against toxic-reduced oxygen species, which are generated as byproducts of many biological oxidations. The generation of oxygen radicals can be further exacerbated during environmental adversity and consequently SOD has been proposed to be important for plant stress tolerance. In plants, three forms of the enzyme exist, as classified by their active site metal ion: copper/zinc, manganese, and iron forms. The distribution of these enzymes has been studied both at the subcellular level and at the phylogenic level. It is only in plants that all three different types of SOD coexist. Their occurrence in the different subcellular compartments of plant cells allows a study of their molecular evolution and the possibility of understanding why three functionally equivalent but structurally different types of SOD have been maintained. Several cDNA sequences that encode the different SODs have recently become available, and the use of molecular techniques have greatly increased our knowledge about this enzyme system and about oxidative stress in plants in general, such that now is an appropriate time to review our current knowledge.     

Superoxide Dismutase in Bacillus popilliae

Vegetative cells of Bacillus popilliae were devoid of catalase but had high levels of superoxide dismutase. This provides further support of a theory that oxygen tolerance by an organism is more dependent on superoxide dismutase than on catalase.     

超氧化物歧化酶(SOD)的分子生物学

本文主要从分子生物学角度概述了超氧化物歧化酶基因的克隆,表达,分子调控,及转基因研究的情况。     

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.     

Assay and Inhibitors of Spinach Superoxide Dismutase

Allosamidin, a product of Streptomyces sp. No 1713, inhibited Bombyx mori chitinase specifically in a competitive way with a Ki o f about 0.1 μm. The effect of allosamidin on chitinases from r Streptomyces griseus and Serratia marcescens was weaker, about 1/500 that on B. mori chitinase. Allosamidin did not inhibit yam chitinase, lysozymes of hen egg-white or human urine, or B. mori α-N-acetyl-d-glucosaminidase. The results suggest that allosamidin is a specific inhibitor of the insect chitinase.     

Mechanism and Atomic Structure of Superoxide Dismutase

The active site Cu ion in Cu,Zn superoxide dismutase is alternately oxidized and reduced during the enzymatic dismutation of superoxide to hydrogen peroxide and molecular oxygen. For oxidized Cu,Zn superoxide dismutase, an atomic structure has been determined for the human enzyme at 2.5 A resolution. The resolution of the bovine enzyme structure has been extended to 1.8 A. Atomic resolution data has been, collected for reduced and inhibitor-bound Cu,Zn superoxide dismutases. and the interpretation of the' electron density difference maps is in progress. The geometry and molecular surfaces of the active sites in these structures, together with biochemical data, suggest a specific model for the enzyme mechanism. Similarities in the active site geometry of the Mn and Fe superoxide dismutases with the Cu.Zn enzyme suggest that dismutation in these enzymes may follow a similar mechanism.     

An Expanded Function for Superoxide Dismutase

α-Hydroxyalkylperoxyl radicals were generated from the primary and secondary alcohols methanol, ethanol and 2-propanol in N₂O/O₂-saturated aqueous solutions by pulse radiolysis. These radicals reduced a ferric iron porphyrin complex, tetrakis-(4-N-methylpyridyl)porphine, with diffusiontontrolled rate constants. The extreme sensitivity of the shift of the Soret absorption band in this reaction was used to determine, by competition kinetics, the reactivity of the peroxyl radicals with different proteins. Only native Cu, Zn-superoxide dismutase and metallothionein showed competitive behavior, with SOD exhibiting rate constants close to the dismutation rate for O₂-. Metallothionein was slower by a factor of 30 with hydroxymethylperoxyl radicals. We propose, that SOD has unique properties of the protein surface in addition to the prosthetic copper site, having possibly evolved as a 'general-purpose radical-scavenging protein'.     

Superoxide Dismutase Therapy for Myocardial Ischemia

Oxygen derived free radicals have been shown to be generated during reperfusion of ischemic myocardium by a variety of approaches including spin trap probes. Three levels of injury have been described for the reperfused heart. Periods of ischemia of only several minutes can trigger lethal arrhythmias on reperfusion. Anti-oxidants including SOD and or catalase, as well as iron chelators reduce the incidence of these arrhythmias in both dog and rat. Xanthine oxidase inhibitors arc equipotent with SOD in this model suggesting that xanthine oxidase is the source of the radicals. Periods of occlusion lasting 10–15 minutes produce a recoverable defect in contractility termed “stunning”. SOD plus catalase has been shown to reduce the incidence of stunning in a variety of models including the xanthine oxidase deficient rabbit. Neither agent on its own seemed to be effective against stunning in either the rabbit or the dog. Stunning is more difficult to demonstrate in the rabbit heart, presumably due to its lack of xanthine oxidase. Periods of ischemia in excess of 20 minutes will result in some irreversible cell death (infarction) with reperfusion. While studies using histochemical methods suggested that SOD plus Catalase given at the time of reper-fusion could limit necrosis in the dog model, histological studies reveal that infarct size was not modified but rather, SOD appears to interfere with the ability of tetrazoliurn to histochemically discriminate between living and dead cells. While PEG SOD with its extended plasma half life was reported to reduce infarct size in the dog. it was unable to protect the reprefused rabbit heart. To date. none of the scavengers have been proven to limit infarction suggesting that free radicals contribute to arrythmias and stunning, but do not. kill cells in the reperfused heart.     

超氧化物歧化酶(SOD)研究进展

环境胁迫使植物细胞中积累大量的活性氧,从而导致蛋白质、膜脂、DNA及其它细胞组分的严重损伤。植物体内有效清除活性氧的酶类包括超氧化物歧化酶(SOD)、抗坏血酸过氧化物酶(APX)、过氧化氢酶(CAT)等,其中研究最深入的是SOD。利用基因工程策略增加这些物质在植物体内的含量,从而获得抗逆转基因植株。     

羊红细胞超氧化物歧化酶的研究

自羊红细胞分离得到一种高等电点的铜.锌-超氧化物歧化酶(Cu.ZnSOD)。其沉降系数(S)为3.23,亚基分子量为16600,等电点为8.50,紫外最大吸收峰位于259nm,酶分子中含有铜和锌,氨基酸组成特点与其它动物来源的Cu.Zn-SOD相同。该酶的比活性为5500U/mg(黄嘌吟氧化酶—细胞色素还原法);对KCN的抑制作用敏感,最适pH值为6。