Dimer destabilization in superoxide dismutase may result in disease-causing properties: Structures of motor neuron disease mutants

Hough, Michael A.; Grossmann, J. Günter; Antonyuk, S.V.; Strange, R.W.; Doucette, P.A.; Rodríguez, Jorge A.; Whitson, Lisa J.; Hart, P. John; Hayward, Lawrence J.; Valentine, Joan Selverstone; Hasnain, S.S.

doi:10.1073/pnas.0305143101

Cited by 203 publications

(226 citation statements)

References 43 publications

(36 reference statements)

Supporting

Mentioning

208

Contrasting

Unclassified

Order By: Relevance

“…X-ray solution scattering data on SOD1 mutant protein (A4V and I113T) also have implied a strikingly different monomer-monomer interface compared with the WT protein (14). Although the SOD1 molecules in all of these structures have the essential disulfide intact, they reveal the ability of mutant proteins to form nonnative protein-protein interactions despite the otherwise WT-like fold.…”

Section: Discussionmentioning

confidence: 99%

“…In contrast, demetallation of SOD1 mutant proteins leads to structural destabilization, the degree of which exhibits the reverse correlation with the mean survival time after ALS diagnosis (11). Several groups, including our own, have proposed that conformational changes of the SOD1 dimer and͞or the dissociation into monomers facilitate protein aggregation (9,(12)(13)(14); in fact, stabilization of SOD1 dimer by small molecules is effective in reducing the protein aggregates in vitro (15). ALS-associated mutations, thus, would inhibit sufficient control of the posttranslational modifications of SOD1, which further leads to the protein destabilization, misfolding, and aggregation.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Disulfide cross-linked protein represents a significant fraction of ALS-associated Cu, Zn-superoxide dismutase aggregates in spinal cords of model mice

Furukawa¹,

Deng

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

195

186

View full text Add to dashboard Cite

Point mutations in Cu, Zn-superoxide dismutase (SOD1) cause a familial form of the neurodegenerative disease amyotrophic lateral sclerosis (ALS). Aggregates of mutant SOD1 proteins are observed in histopathology and are invoked in several proposed mechanisms for motor neuronal death; however, the significant stability and activity of the mature mutant proteins are not readily explained in such models. Recent biochemical studies suggest that it is the immature disulfide-reduced forms of the familial ALS mutant SOD1 proteins that play a critical role; these forms tend to misfold, oligomerize, and readily undergo incorrect disulfide formation upon mild oxidative stress in vitro. Here we provide physiological support for this mechanism of aggregate formation and show that a significant fraction of the insoluble SOD1 aggregates in spinal cord of the ALS-model transgenic mice contain multimers cross-linked via intermolecular disulfide bonds. These insoluble disulfide-linked SOD1 multimers are found only in the spinal cord of symptomatic transgenic animals, are not observed in unafflicted tissue such as brain cortex and liver, and can incorporate WT SOD1 protein. The findings provide a biochemical basis for a pathological hallmark of this disease; namely, incorrect disulfide cross-linking of the immature, misfolded mutant proteins leads to insoluble aggregates. disulfide bond ͉ protein aggregation ͉ oxidative stress ͉ neurodegnerative disease A myotrophic lateral sclerosis (ALS) is one of the most common adult-onset neurodegenerative diseases, and, in 1993, several mutations in the Cu, Zn-superoxide dismutase (SOD1) gene were identified as a cause of a subset of familial ALS (fALS) (1, 2). Since that time, Ͼ100 types of SOD1 mutations have been linked with fALS; however, its molecular mechanism remains unresolved. SOD1, which is a homodimer with a copper and zinc ion in each subunit, functions as an antioxidant enzyme by converting superoxide radical to oxygen and hydrogen peroxide (3). Some of the SOD1 mutant proteins have comparable levels of dismutase activity, and the SOD1-knockout mouse does not develop ALS-like motor neuron symptoms (4), leading to the idea that SOD1-mediated toxicity in fALS is due to a new activity acquired by mutations in the SOD1 gene. One of the proposed toxic functions in the SOD1 mutant proteins is an aberrant copper chemistry. A subset of ALS mutations in SOD1 can lead to catalytic nitration of tyrosine residues and peroxide or superoxide production at the copper site (5); however, the copper-toxicity model is difficult to reconcile with the fact that ALS symptoms still are observed in the transgenic mouse expressing SOD1 proteins in which all four copper ligands are mutated (6). Although some copper chelators can reduce the toxicity of mutant proteins in transgenic ALS model mice (7), a role for aberrant copper chemistry in the disease has not been widely addressed.Another hypothesis for a gain of toxic function is that the mutations in SOD1 facilitate protein aggregation (8). An enzyma...

show abstract

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Disulfide cross-linked protein represents a significant fraction of ALS-associated Cu, Zn-superoxide dismutase aggregates in spinal cords of model mice

Furukawa¹,

Deng

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

195

186

View full text Add to dashboard Cite

show abstract

“…Furthermore, the additional secondary structure in its C-terminal region suggests an interference with proper dimerization, which might lead to conformational changes in its catalytically active region, resulting in an inactive and/or instable form. Similarly, a growing body of evidence suggests that a subset of mutations located close to the dimeric interface can lead to a major destabilization of mouse Cu/Zn-SOD1 (Hough et al, 2004;Hornberg et al, 2007). Additional types of possible regulation at the posttranscriptional level (e.g.…”

Section: Resultsmentioning

confidence: 99%

Alternative Splicing Studies of the Reactive Oxygen Species Gene Network inPopulusReveal Two Isoforms of High-Isoelectric-Point Superoxide Dismutase

Srivastava

Chibani

et al. 2009

Plant Physiology

View full text Add to dashboard Cite

Recent evidence has shown that alternative splicing (AS) is widely involved in the regulation of gene expression, substantially extending the diversity of numerous proteins. In this study, a subset of expressed sequence tags representing members of the reactive oxygen species gene network was selected from the PopulusDB database to investigate AS mechanisms in Populus. Examples of all known types of AS were detected, but intron retention was the most common. Interestingly, the closest Arabidopsis (Arabidopsis thaliana) homologs of half of the AS genes identified in Populus are not reportedly alternatively spliced. Two genes encoding the protein of most interest in our study (high-isoelectric-point superoxide dismutase [hipI-SOD]) have been found in black cottonwood (Populus trichocarpa), designated PthipI-SODC1 and PthipI-SODC2. Analysis of the expressed sequence tag libraries has indicated the presence of two transcripts of PthipI-SODC1 (hipI-SODC1b and hipISODC1s). Alignment of these sequences with the PthipI-SODC1 gene showed that hipI-SODC1b was 69 bp longer than hipISODC1s due to an AS event involving the use of an alternative donor splice site in the sixth intron. Transcript analysis showed that the splice variant hipI-SODC1b was differentially expressed, being clearly expressed in cambial and xylem, but not phloem, regions. In addition, immunolocalization and mass spectrometric data confirmed the presence of hipI-SOD proteins in vascular tissue. The functionalities of the spliced gene products were assessed by expressing recombinant hipI-SOD proteins and in vitro SOD activity assays.

show abstract

“…Indeed, variant proteins, such as the G85R SOD1 used in this study, are found as monomers in vivo (29)(30)(31). A number of modifications, including loss of Cu or Zn (32), cleavage of the native, intramolecular disulfide (33), oxidation (13,34), and fALS-associated mutation (35), predispose the SOD1 dimer to dissociate. X-ray crystal structures of both A4V, and to a lesser extent I113T (35); yeast two hybrid analysis of H46R, A4V, and H48Q (36); dissociation of G85R, G93R, E100G, and I113T by chaotrophs (37); and molecular dynamics simulations (38,39) are all consistent with this hypothesis.…”

mentioning

confidence: 99%

Strategies for stabilizing superoxide dismutase (SOD1), the protein destabilized in the most common form of familial amyotrophic lateral sclerosis

Auclair

Boggio

Petsko

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

Amyotrophic lateral sclerosis (ALS) is a disorder characterized by the death of both upper and lower motor neurons and by 3-to 5-yr median survival postdiagnosis. The only US Food and Drug Administration-approved drug for the treatment of ALS, Riluzole, has at best, moderate effect on patient survival and quality of life; therefore innovative approaches are needed to combat neurodegenerative disease. Some familial forms of ALS (fALS) have been linked to mutations in the Cu/Zn superoxide dismutase (SOD1). The dominant inheritance of mutant SOD1 and lack of symptoms in knockout mice suggest a "gain of toxic function" as opposed to a loss of function. A prevailing hypothesis for the mechanism of the toxicity of fALS-SOD1 variants, or the gain of toxic function, involves dimer destabilization and dissociation as an early step in SOD1 aggregation. Therefore, stabilizing the SOD1 dimer, thus preventing aggregation, is a potential therapeutic strategy. Here, we report a strategy in which we chemically cross-link the SOD1 dimer using two adjacent cysteine residues on each respective monomer (Cys111). Stabilization, measured as an increase in melting temperature, of ∼20°C and ∼45°C was observed for two mutants, G93A and G85R, respectively. This stabilization is the largest for SOD1, and to the best of our knowledge, for any disease-related protein. In addition, chemical cross-linking conferred activity upon G85R, an otherwise inactive mutant. These results demonstrate that targeting these cysteine residues is an important new strategy for development of ALS therapies.mass spectrometry | thiol-disulfide I nnovative approaches are needed to combat neurodegenerative disease, among the most serious of which is amyotrophic lateral sclerosis (ALS), a disorder characterized by the death of both upper and lower motor neurons and by 3-to -5-yr median survival postdiagnosis. The only US Food and Drug Administrationapproved drug for the treatment of ALS, Riluzole, has at best, moderate effect on patient survival and quality of life (1-3). Although the causes of sporadic neurodegenerative diseases remain a mystery, mutations causing familial forms of many of these diseases (e.g., Alzheimer's, Parkinson, and ALS) are known. For example, mutations in the gene encoding Cu/Zn superoxide dismutase (SOD1) are responsible for ∼20% of the familial ALS cases (fALS) and 2% of all ALS (4, 5). Two such mutations are G93A, which maintains wild-type-like enzymatic activity, and the metal-deficient G85R, which is essentially inactive. Posttranslational modifications of proteins involved in familial diseases have been invoked in the etiology of the corresponding sporadic diseases, for example, alpha-synuclein (6) and Parkin (7) modification in Parkinson, Abeta (8) and tau (9) modification in Alzheimer's, and TDP43 (10) and SOD1 (11-14) modification in ALS. The hope, therefore, is that strategies for treating familial diseases may translate to at least a subset of sporadic diseases.Both dominant inheritance of mutant SOD1 (15) and lack of symptoms i...

show abstract

Dimer destabilization in superoxide dismutase may result in disease-causing properties: Structures of motor neuron disease mutants

Cited by 203 publications

References 43 publications

Disulfide cross-linked protein represents a significant fraction of ALS-associated Cu, Zn-superoxide dismutase aggregates in spinal cords of model mice

Disulfide cross-linked protein represents a significant fraction of ALS-associated Cu, Zn-superoxide dismutase aggregates in spinal cords of model mice

Alternative Splicing Studies of the Reactive Oxygen Species Gene Network inPopulusReveal Two Isoforms of High-Isoelectric-Point Superoxide Dismutase

Strategies for stabilizing superoxide dismutase (SOD1), the protein destabilized in the most common form of familial amyotrophic lateral sclerosis

Contact Info

Product

Resources

About