Over 100 point mutations in the Cu/Zn superoxide dismutase (SOD) gene have been linked to the neurodegenerative disease amyotrophic lateral sclerosis (ALS). Exactly why these mutations cause ALS is unclear, but abounding explanations fall into two main camps-those that hinge on mutants being catalytically active, and those that do not. A report in today's PNAS from researchers at Umea University, Sweden, strongly supports the latter view.

Senior author Mikael Oliveberg and colleagues characterized ALS-associated SOD mutants containing single amino-acid substitutions in various parts of the protein. The authors found that one thing all mutants had in common is that their apo protein, i.e., the protein alone without its active-site metal ions, is less stable than wild-type apo protein. First author Mikael Lindberg et al. determined this by denaturing the proteins with either guanidium chloride or urea.

The authors found that substituting alanine for glycine at position 93, for example, resulted in complete denaturation of the polypeptide at a concentration of two molar urea, whereas four molar is needed to denature wild-type apo protein. Mutating N-terminal amino acids (valine for alanine at position 4, and phenylalanine for cysteine at postion 6), which are buried in the hydrophobic core of the protein, had even more drastic effects. Interestingly, substituting alanine for cysteine at position 6, a mutation that is not associated with ALS but is found in other, presumably healthy organisms, has no effect on the stability of the apo protein.

The results suggest that destabilization of the immature protein may be a contributing factor to ALS progression. This supports recent data indicating that copper loading of SOD is not required for manifestation of the disease (see related new item). It is also noteworthy that the least stable mutants (i.e., A4V, and C6F) are also associated with faster progression of ALS.—Tom Fagan

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  1. First of all I must admit that I am not an expert on the dynamics of SOD protein folding. In this paper, Lindberg et al. provide data that suggests the involvement of Apo-SOD in ALS disease pathogenesis. It has been known for a long time that the development of the disease in familial SOD1 mutation is not dependent on SOD enzymatic activity and the mutation is a dominant gain of function. Alterations in expression of wild-type SOD enzyme by over-expression or targeted deletion of SOD show no effect on the disease course of mutant SOD1 mice. More recently it was shown that substantial reduction in copper binding activity by deletion of CCS does not alter disease course in the SOD1 mice (see ARF related news story), suggesting again that SOD enzymatic activity may not be involved in the disease process. This paper supports these earlier observations and lends more credence to the idea that SOD1 mutation causes a protein conformation disease. The generation of high molecular weight isoforms of SOD, SOD1 inclusions and alterations in SOD protein folding in general point in that direction.

    However, the suggestion by the authors that it is Apo form of the protein that is toxic and the suggestion that instability of Apo-SOD correlates with clinical course of the disease is premature. Specifically, the comparison of the D90A and A4V phenotype in humans to the stability of their respective Apo-SOD is misleading. While the A4V is predominantly a dominant mutation, the D90A mutation is mostly recessive. The dominant D90A mutation identified in some patients, however, causes a rapidly progressing disease and is a more appropriate comparison to the A4V mutation.

    Although the SOD protein is toxic, it is still not clear how the toxicity is produced. SOD protein is expressed throughout development, but it is still not clear why it takes decades to produce the disease. Recently, Neil Cashman has suggested a model similar to the prion model that involved isomorphic recruitment of SOD1 in ALS, which supports a seeding hypothesis. Once seeded, it is not clear which supramolecular isoform of SOD1 is toxic and how they generate toxicity. Apo enzyme, holoenzyme and/or both may be involved in seeding and formation of the supramolecular isoforms. Future experiments to address these questions will provide a good handle on understanding ALS and also diseases such as Alzheimer’s disease, CJD, cystic fibrosis, Huntington’s disease, all of which fall under this new class of protein conformation disorders.

References

News Citations

  1. Disputing the Dismuting—What Is the Real Role of SOD in ALS?

Further Reading

No Available Further Reading

Primary Papers

  1. . Common denominator of Cu/Zn superoxide dismutase mutants associated with amyotrophic lateral sclerosis: decreased stability of the apo state. Proc Natl Acad Sci U S A. 2002 Dec 24;99(26):16607-12. PubMed.