共查询到20条相似文献,搜索用时 15 毫秒
1.
Gunther MR Vangilder R Fang J Beattie DS 《Archives of biochemistry and biophysics》2004,431(2):207-214
Strains of Saccharomyces cerevisiae that express either the wild type or the amyotrophic lateral sclerosis-associated mutant human copper-zinc superoxide dismutase (SOD1) proteins A4V and G93A, respectively, in a yeast SOD1-deficient parent strain were used to investigate the hypothesis that expression of a mutant SOD1 protein causes deficient mitochondrial electron transport as a possible mechanism for disease induction. Mitochondria isolated from the wild type SOD1-expressing yeast were identical to mitochondria from the parent strain in heme content and activities of complexes II, III, and IV. Mitochondria isolated from the A4V-expressing yeast had decreased rates of electron transport in complexes II+III, III, and IV and corresponding decreases in hemes b, c-c1, and a-a3 content compared to mitochondria from wild type human SOD1-expressing yeast. Mitochondria isolated from G93A-expressing yeast had decreased rates of electron transport in complex IV and probably in complex II with a corresponding decrease in heme a-a3 content. These results suggest that mutant SOD1-expression causes defective electron transport complex assembly and that the yeast system will provide an excellent model for the study of the mechanism of mutant SOD1-induced mitochondrial electron transport defects. 相似文献
2.
Mutations in Cu/Zn superoxide dismutase (SOD) are involved in some cases of familial amyotrophic lateral sclerosis, and it appears that misfolding and aggregation, perhaps mediated by abnormal binding or loss of copper (Cu) and/or zinc (Zn), may play a pathological role. It is known that the absence of both metals kinetically destabilizes wild type and mutant SOD leading to a 60-fold increase in their rate of unfolding. Here, the individual contributions of Cu and Zn to the kinetic stability of SOD were investigated, and the results show that Cu plays a greater role. Thus, the deficiency of Cu or Zn, especially the former, will compromise the kinetic stability of SOD, thereby increasing the probability that pathogenic mutants and even the WT protein may misfold and self-assemble into toxic species. 相似文献
3.
T. J. Lyons A. Nersissian H. Huang H. Yeom C. R. Nishida J. A. Graden E. B. Gralla J. S. Valentine 《Journal of biological inorganic chemistry》2000,5(2):189-203
We have investigated factors that influence the properties of the zinc binding site in yeast copper-zinc superoxide dismutase
(CuZnSOD). The properties of yeast CuZnSOD are essentially invariant from pH 5 to pH 9. However, below this pH range there
is a change in the nature of the zinc binding site which can be interpreted as either (1) a change in metal binding affinity
from strong to weak, (2) the expulsion of the metal bound at this site, or (3) a transition from a normal distorted tetrahedral
ligand orientation to a more symmetric arrangement of ligands. This change is strongly reminiscent of a similar pH-induced
transition seen for the bovine protein and, based on the data presented herein, is proposed to be a property that is conserved
among CuZnSODs. The transition demonstrated for the yeast protein is not only sensitive to the pH of the buffering solution
but also to the occupancy and redox status of the adjacent copper binding site. Furthermore, we have investigated the effect
of single site mutations on the pH- and redox-sensitivity of Co2+ binding at the zinc site. Each of the mutants H46R, H48Q, H63A, H63E, H80C, G85R, and D83H is capable of binding Co2+ to a zinc site with a distorted tetrahedral geometry similar to that of wild-type. However, they do so only if Cu+ is bound at the copper site or if the pH in raised to near physiological levels, indicating that the change at the zinc binding
site seen in the wild-type is conserved in the mutants, albeit with an altered pK
a. The mutants H71C and D83A did not bind Co2+ in a wild-type-like fashion under any of the conditions tested. This study reveals that the zinc binding site is exquisitely
sensitive to changes in the protein environment. Since three of the mutant yeast proteins investigated here contain mutations
analogous to those that cause ALS (amyotrophic lateral sclerosis) in humans, this finding implicates improper metal binding
as a mechanism by which CuZnSOD mutants exert their toxic gain of function.
Received: 17 September 1999 / Accepted: 8 December 1999 相似文献
4.
Paul C. Trippier Radhia Benmohamed Donald R. Kirsch Richard B. Silverman 《Bioorganic & medicinal chemistry letters》2012,22(21):6647-6650
Amyotrophic lateral sclerosis (ALS) is a debilitating and fatal neurodegenerative disease. Although the cause remains unknown, misfolded protein aggregates are seen in neurons of sporadic ALS patients, and familial ALS mutations, including mutations in superoxide dismutase 1 (SOD1), produce proteins with an increased propensity to misfold and aggregate. A structure activity relationship of a lead scaffold exhibiting neuroprotective activity in a G93A-SOD1 mouse model for ALS has been further investigated in a model PC12 cellular assay. Synthesis of biotinylated probes at the N1 nitrogen of the pyrazolone ring gave compounds (5d–e) that retained activity within 10-fold of the proton-bearing lead compound (5a) and were equipotent with a sterically less cumbersome N1-methyl substituted analogue (5b). However, when methyl substitution was introduced at N1 and N2 of the pyrazolone ring, the compound was inactive (5c). These data led us to investigate further the pharmacophoric nature of the pyrazolone unit. A range of N1 substitutions were tolerated, leading to the identification of an N1-benzyl substituted pyrazolone (5m), equipotent with 5a. Substitution at N2 or excision of N2, however, removed all activity. Therefore, the hydrogen bond donating ability of the N2–H of the pyrazolone ring appears to be a critical part of the structure, which will influence further analogue synthesis. 相似文献
5.
Mutations in superoxide dismutase 1 (SOD1) cause some forms of familial amyotrophic lateral sclerosis (fALS). Affected tissues of patients and transgenic mouse models of the disease accumulate misfolded and aggregated forms of the mutant protein. In the present study we have identified specific sequences in human SOD1 that modulate the aggregation of fALS mutant proteins. From our study of a panel of mutant proteins, we identify two sequence elements in human SOD1 (residues 42-50 and 109-123) that are critical in modulating the aggregation of the protein. These sequences are components of the 4th and 7th β-strands of the protein, and in the native structure are normally juxtaposed as elements of the core β-barrel. Our data suggest that some type of intermolecular interaction between these elements may occur in promoting mutant SOD1 aggregation. 相似文献
6.
Rhoads TW Lopez NI Zollinger DR Morré JT Arbogast BL Maier CS DeNoyer L Beckman JS 《Analytical biochemistry》2011,(1):52-58
Metals are key cofactors for many proteins, yet quantifying the metals bound to specific proteins is a persistent challenge in vivo. We have developed a rapid and sensitive method using electrospray ionization mass spectrometry to measure Cu,Zn superoxide dismutase (SOD1) directly from the spinal cord of SOD1-overexpressing transgenic rats. Metal dyshomeostasis has been implicated in motor neuron death in amyotrophic lateral sclerosis (ALS). Using the assay, SOD1 was directly measured from 100 μg of spinal cord, allowing for anatomical quantitation of apo, metal-deficient, and holo SOD1. SOD1 was bound on a C4 Ziptip that served as a disposable column, removing interference by physiological salts and lipids. SOD1 was eluted with 30% acetonitrile plus 100 μM formic acid to provide sufficient hydrogen ions to ionize the protein without dislodging metals. SOD1 was quantified by including bovine SOD1 as an internal standard. SOD1 could be measured in subpicomole amounts and resolved to within 2 Da of the predicted parent mass. The methods can be adapted to quantify modifications to other proteins in vivo that can be resolved by mass spectrometry. 相似文献
7.
为探索简便实用纯化SOD的工艺路线,以人或猪血红细胞溶血上清液,经铜胺中空纤维透析器(分子量截留值为15kD)透析和超滤,收集分子量大于15.0kD的物质,再加热60℃10min,离心取上清即得。Cu、ZnSOD和MnSOD分子量分别为32.0kD和80.0kD。人血和猪血纯化的SOD总收率分别为88.2%和89.2%,比活性分别为17429U/mg和18228U/mg。工艺简便实用,适于工业纯化生产。 相似文献
8.
《Bioorganic & medicinal chemistry letters》2014,24(6):1532-1537
The treatment of neurodegenerative diseases is difficult because of multiple etiologies and the interplay of genetics and environment as precipitating factors. In the case of amyotrophic lateral sclerosis (ALS), we have knowledge of a handful of genes that cause disease when mutated. However, drugs to counteract the effect of genetic mutations have not yet been found. One of the causative genes, Cu, Zn-superoxide dismutase (SOD1) is responsible for about 10–15% of the genetically linked autosomal dominant disease. Our rationale was that compounds that reduce expression of the mutant protein would be beneficial to slow onset and/or disease progression. We screened candidate compounds using a cell-based in vitro assay for those that reduce mutant SOD1 (G93A) protein expression. This led to the discovery of 2-[3-iodophenyl)methylsulfanyl]-5pyridin-4-yl-1,3,4-oxadiazole, a known protein kinase inhibitor that decreases G93A-SOD1 expression in vitro and in the brain and spinal cord in vivo. However, this compound has a biphasic dose response curve and a likely toxophore which limit its therapeutic window for chronic disease such as ALS. Therefore, we designed and tested a focused library of analogs for their ability to decrease SOD1 expression in vitro. This exercise resulted in the identification of a lead compound with improved drug-like characteristics and activity. Development of small molecules that reduce the expression of etiologically relevant toxic proteins is a strategy that may also be extended to familial ALS linked to gain of function mutations in other genes. 相似文献
9.
10.
Elevated cortical extracellular fluid glutamate in transgenic mice expressing human mutant (G93A) Cu/Zn superoxide dismutase 总被引:6,自引:0,他引:6
Alexander GM Deitch JS Seeburger JL Del Valle L Heiman-Patterson TD 《Journal of neurochemistry》2000,74(4):1666-1673
Transgenic mice expressing a mutated (G93A) human Cu/Zn superoxide dismutase (SOD1) develop motor neuron pathology and clinical symptoms similar to those seen in patients with amyotrophic lateral sclerosis. Loss of motor neurons is most prominent in lumbar, followed by cervical cord and then brainstem. No significant cell death has been reported in motor cortex. The integrity of the cortical glutamate reuptake systems was evaluated using intracerebral microdialysis and western immunoblot assays for the glutamate transporters GLT-1, GLAST, and EAAC1. The basal extracellular fluid levels of aspartate, glutamate, glutamine, 3,4-dihydroxyphenylacetic acid, and 5-hydroxyindole-3-acetic acid were evaluated by HPLC. The extraction fraction of L-3H]glutamate, corrected with [14C]mannitol, was also evaluated. GLT-1, EAAC1, and GLAST protein levels were determined by semiquantitative chemiluminescence immunoblot of proteins from membrane-enriched fractions. The relative optical density of film was translated into relative protein level by comparison with a standard control mouse. The SOD1 mutant mice demonstrated a significant (p < 0.05) increase in basal levels of extracellular aspartate and glutamate. In addition, when the glutamate extraction fraction was challenged with exogenous unlabeled glutamate (500 microM) by reversed microdialysis, the glutamate extraction fraction in the mutant SOD1 mice was decreased significantly from control levels. The SOD1 mutant mice demonstrated no difference in the cortical protein levels of the glutamate transporter subtypes. This study demonstrates that in areas of no visible pathology and no loss of glutamate transporter proteins, SOD1 mutant mice have elevated extracellular fluid aspartate and glutamate levels and a decreased capacity to clear glutamate from the extracellular space. 相似文献
11.
Mutant SOD1 linked to familial amyotrophic lateral sclerosis,but not wild-type SOD1, induces ER stress in COS7 cells and transgenic mice 总被引:6,自引:0,他引:6
Tobisawa S Hozumi Y Arawaka S Koyama S Wada M Nagai M Aoki M Itoyama Y Goto K Kato T 《Biochemical and biophysical research communications》2003,303(2):496-503
Mutations in a Cu, Zn-superoxide dismutase (SOD1) cause motor neuron death in human familial amyotrophic lateral sclerosis (FALS) and its mouse model, suggesting that mutant SOD1 has a toxic effect on motor neurons. However, the question of how the toxic function is gained has not been answered. Here, we report that the mutant SOD1s linked to FALS, but not wild-type SOD1, aggregated in association with the endoplasmic reticulum (ER) and induced ER stress in the cDNA-transfected COS7 cells. These cells showed an aberrant intracellular localization of mitochondria and microtubules, which might lead to a functional disturbance of the cells. Motor neurons of the spinal cord in transgenic mice with a FALS-linked mutant SOD1 also showed a marked increase of GRP78/BiP, an ER-resident chaperone, just before the onset of motor symptoms. These data suggest that ER stress is involved in the pathogenesis of FALS with an SOD1 mutation. 相似文献
12.
13.
Chung J Yang H de Beus MD Ryu CY Cho K Colón W 《Biochemical and biophysical research communications》2003,312(4):873-876
Mutations in Cu/Zn superoxide dismutase (SOD) are associated with familial amyotrophic lateral sclerosis (FALS), a neurodegenerative disease that is characterized by the selective death of motor neurons. Despite the genetic association made between the protein and the disease, the mechanism by which the mutant SOD proteins become toxic is still a mystery. Using wild-type SOD and three pathogenic mutants (A4V, G37R, and G85R), we show that the copper-induced oxidation of metal-depleted SOD causes its in vitro aggregation into pore-like structures, as determined by atomic force microscopy. Because toxic pores have been recently implicated in the pathogenic mechanism of other neurodegenerative diseases, these results raise the possibility that the aberrant self-assembly of oxidatively damaged SOD mutants into toxic oligomers or pores may have a pathological role in FALS. 相似文献
14.
A supramolecular bifunctional artificial enzyme with superoxide dismutase and glutathione peroxidase activities 总被引:1,自引:0,他引:1
Shuangjiang Yu 《Bioorganic chemistry》2010,38(4):159-827
For constructing a bifunctional antioxidative enzyme with both superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities, a supramolecular artificial enzyme was successfully constructed by the self-assembly of the Mn(III)meso-tetra[1-(1-adamantyl methyl ketone)-4-pyridyl] porphyrin (MnTPyP-M-Ad) and cyclodextrin-based telluronic acid (2-CD-TeO3H) through host-guest interaction in aqueous solution. The self-assembly of the adamantyl moieties of Mn(III) porphyrin and the β-CD cavities of 2-CD-TeO3H was demonstrated by the NMR spectra. In this supramolecular enzyme model, the Mn(III) porphyrin center acted as an efficient active site of SOD and tellurol moiety endowed GPx activity. The SOD-like activity (IC50) of the new catalyst was found to be 0.116 μM and equals to 2.56% of the activity of the native SOD. Besides this, supramolecular enzyme model also showed a high GPx activity, and a remarkable rate enhancement of 27-fold compared to the well-known GPx mimic ebselen was observed. More importantly, the supramolecular artificial enzyme showed good thermal stability. 相似文献
15.
超氧化物岐化酶在医学领域的研究现状 总被引:2,自引:0,他引:2
超氧化物歧化酶(SOD)是生物体内清除超氧阴离子自由基(O2-.)的一种重要金属酶,具有重要的生理功能,在医药、食品等方面有广泛的应用前景,是目前医学、分子生物学领域研究的热点之一。现对SOD的分类、分布、结构、性质、在相关疾病中的研究进展、临床应用等方面做一综述。 相似文献
16.
Variable metallation of human superoxide dismutase: atomic resolution crystal structures of Cu-Zn, Zn-Zn and as-isolated wild-type enzymes 总被引:1,自引:0,他引:1
Strange RW Antonyuk SV Hough MA Doucette PA Valentine JS Hasnain SS 《Journal of molecular biology》2006,356(5):1152-1162
Human Cu-Zn superoxide dismutase (SOD1) protects cells from the effects of oxidative stress. Mutations in SOD1 are linked to the familial form of amyotrophic lateral sclerosis. Several hypotheses for their toxicity involve the mis-metallation of the enzyme. We present atomic-resolution crystal structures and biophysical data for human SOD1 in three metallation states: Zn-Zn, Cu-Zn and as-isolated. These data represent the first atomic-resolution structures for human SOD1, the first structure of a reduced SOD1, and the first structure of a fully Zn-substituted SOD1 enzyme. Recombinantly expressed as-isolated SOD1 contains a mixture of Zn and Cu at the Cu-binding site. The Zn-Zn structure appears to be at least as stable as the correctly (Cu-Zn) metallated enzyme. These data raise the possibility that in a cellular environment with low availability of free copper, Zn-Zn may be the preferred metallation state of SOD1 prior to its interaction with the copper chaperone. 相似文献
17.
Increased affinity for copper mediated by cysteine 111 in forms of mutant superoxide dismutase 1 linked to amyotrophic lateral sclerosis 总被引:1,自引:0,他引:1
Watanabe S Nagano S Duce J Kiaei M Li QX Tucker SM Tiwari A Brown RH Beal MF Hayward LJ Culotta VC Yoshihara S Sakoda S Bush AI 《Free radical biology & medicine》2007,42(10):1534-1542
Mutations in Cu,Zn-superoxide dismutase (SOD1) cause familial amyotrophic lateral sclerosis (ALS). It has been proposed that neuronal cell death might occur due to inappropriately increased Cu interaction with mutant SOD1. Using Cu immobilized metal-affinity chromatography (IMAC), we showed that mutant SOD1 (A4V, G85R, and G93A) expressed in transfected COS7 cells, transgenic mouse spinal cord tissue, and transformed yeast possessed higher affinity for Cu than wild-type SOD1. Serine substitution for cysteine at the Cys111 residue in mutant SOD1 abolished the Cu interaction on IMAC. C111S substitution reversed the accelerated degradation of mutant SOD1 in transfected cells, suggesting that the Cys111 residue is critical for the stability of mutant SOD1. Aberrant Cu binding at the Cys111 residue may be a significant factor in altering mutant SOD1 behavior and may explain the benefit of controlling Cu access to mutant SOD1 in models of familial ALS. 相似文献
18.
Yuriy A. Vladimirov Elena A. Gorbatenkova Nikolai V. Paramonov Ofelia A. Azizova 《Free radical biology & medicine》1988,5(5-6):281-286
The effects of helium-neon laser (HNL) on activity, absorption spectra, and ESR signals of superoxide dismutase (SOD; E Cul. 15.1.1) from bovine erythrocytes in acid medium were investigated. It was found that incubation during 2 hours at pH 5.9 led to inactivation of the enzyme. The subsequent illumination of SOD by HNL brought about the enzyme reactivation. Both absorption and ESR spectra were changed after incubation at pH 5.9 and restored after laser irradiation. In a model system, copper-histidine complex, absorption maximum was shifted from 632–633 nm at pH 5.8 to 639–640 nm at pH 8.5–9.0. The similar shift of the maximum was observed after illumination by HNL at pH 5.8. It may be postulated that the photoreactivation of SOD consists essentially in deprotonation of His-61 residue in the enzyme active site and subsequent recovery of imidasol bridge between copper and zinc which had been destroyed at low pH.
Since many other enzymes possess similar histidine-copper structures in their active sites, one may expect diverse effects of red (laser) light on the enzyme activity. Heme-containing enzyme, catalase was also found to be photoreactivated by HNL after inactivation at pH 6.0. 相似文献
19.
Piacham T Isarankura-Na-Ayudhya C Nantasenamat C Yainoy S Ye L Bülow L Prachayasittikul V 《Biochemical and biophysical research communications》2006,341(4):925-930
Superoxide dismutase (SOD) activities of various metallobacitracin complexes were evaluated using the riboflavin-methionine-nitro blue tetrazolium assay. The radical scavenging activity of various metallobacitracin complexes was shown to be higher than those of the negative controls, e.g., free transition metal ions and metal-free bacitracin. The SOD activity of the complex was found to be in the order of Mn(II)>Cu(II)>Co(II)>Ni(II). Furthermore, the effect of bacitracin and their complexation to metals on various microorganisms was assessed by antibiotic susceptibility testing. Moreover, molecular modeling and quantum chemical calculation of the metallobacitracin complex was performed to evaluate the correlation of electrostatic charge of transition metal ions on the SOD activity. 相似文献
20.
Mercedes Prudencio Armando Durazo‡ Julian P. Whitelegge† David R. Borchelt 《Journal of neurochemistry》2009,108(4):1009-1018
Mutations in superoxide dismutase 1 (SOD1, EC 1.15.1.1) cause familial amyotrophic lateral sclerosis; with aggregated forms of mutant protein accumulating in spinal cord tissues of transgenic mouse models and human patients. Mice over-expressing wild-type human SOD1 (WT hSOD1) do not develop amyotrophic lateral sclerosis-like disease, but co-expression of WT enzyme at high levels with mutant SOD1 accelerates the onset of motor neuron disease compared with mice expressing mutant hSOD1 alone. Spinal cords of mice expressing both proteins contain aggregated forms of mutant protein and, in some cases, evidence of co-aggregation of WT hSOD1 enzyme. In the present study, we used a cell culture model of mutant SOD1 aggregation to examine how the presence of WT SOD1 affects mutant protein aggregation, finding that co-expression of WT SOD1, hSOD1 or mouse SOD1, delayed the formation of mutant hSOD1 aggregates; in essence appearing to slow the aggregation rate. In some combinations of WT and mutant hSOD1 co-expression, the aggregates that did eventually form appeared to contain WT hSOD1 protein. However, WT mouse SOD1 did not co-aggregate with mutant hSOD1 despite displaying a similar ability to slow mutant hSOD1 aggregation. Together, these studies indicate that WT SOD1 (human or mouse), when expressed at levels equivalent to the mutant protein, modulates the aggregation of mutant SOD1. 相似文献