Frustrated ALS Enzyme: SOD1 Sacrifices Structural Stability for Function

For superoxide dismutase 1 (SOD1), the metalloenzyme implicated in some forms of familial amyotrophic lateral sclerosis (ALS), the very features that allow the protein to do its job may make it prone to form abnormal aggregates. So conclude the authors of a structural biology study published online in PNAS this week. In a tour de force encompassing folding assays, NMR, calorimetry, and protein crystallography, the scientists in the laboratory of Mikael Oliveberg at Stockholm University in Sweden analyzed the properties of SOD1 when they took away the residues that normally bind a zinc ion. They determined that the zinc is present to stabilize the molecule’s otherwise floppy catalytic loops.

To mature, SOD1 acquires copper and zinc cofactors, forms intramolecular disulfide bonds, and dimerizes. Other studies have pointed to a role for the metal ions in stabilizing SOD1 structure; for example, the Apo form of the protein is more likely to aggregate (see ARF related news story and Banci et al., 2009). In the current study, joint first authors Anna Nordlund and Lina Leinartait and colleagues found that when they removed the zinc-binding site from the protein, it was more prone to aggregation. There is probably now enough evidence to consider these findings irrefutable, wrote Jeffrey Agar of Brandeis University in Waltham, Massachusetts, in an e-mail to ARF. Agar was not involved in the Stockholm study.

SOD1’s copper is functionally required for the enzyme to neutralize free radicals, but the zinc appears to play a more structural role, Oliveberg wrote to ARF. To isolate the importance of the zinc, the scientists knocked out its binding site, mutating the ion’s ligands H63, H71, H80, and D83 to serine. The mutant protein still folded like the wild-type—but then held its structure more stably than the normal protein. The mutant SOD1 was less likely to unfold, and able to withstand higher concentrations of urea, than the native protein. Thus, the residues needed to bind zinc in SOD1’s mature form actually make it less stable when the protein is still immature.

The mutant protein still picked up a zinc ion, but did so with the sites that usually bind copper, which couldn’t position it properly. X-ray crystallography and NMR showed that the misaligned metal ion forced the protein’s catalytic loops to swing outward in a disordered fashion. Those wildly swaying loops, then, were more likely to reach out and bind another SOD1 monomer, potentially seeding the aggregation process. In contrast, the same loops in metal-bound SOD1 were rigid. Oliveberg concluded that the zinc is present to keep those loops tied down.

Oliveberg suggested that SOD1’s inclination toward aggregation is the result of an evolutionary tug-of-war between enzyme function and protein stability. Like the long neck of a giraffe can sometimes cause problems, the long functional loops of SOD1 compromise the protein’s structural integrity, he wrote. Misfolded intermediates accumulate because the functional parts of the protein pull in the wrong direction. Studies show that a few other proteins, such as the trefoil proteins, have a similar folding dilemma—the scientific term is frustrated (Capraro et al., 2008). SOD1 is the first such protein to be implicated in disease, Oliveberg wrote.

SOD1 is a highly conserved, ancient protein, and may be a sort of molecular dinosaur, Oliveberg wrote. It may now be so specialized that any mutation throws the protein off balance. Perhaps it has painted itself into an evolutionary cul-de-sac: the function is essential, but puts a lot of strain’ on the structure, so mutations are difficult to tolerate. That would help explain why more than 100 different mutations in SOD1 have been linked to inherited ALS.

Reference:
Nordlund A, Leinartait L, Saraboji K, Aisenbrey C, Grbner G, Zetterstrm P, Danielsson J, Logan DT, Oliveberg M. Functional features cause misfolding of the ALS-provoking enzyme SOD1. PNAS Early Edition.


To view commentaries, primary articles and linked stories, go to the original posting on Alzforum.org here.

Copyright © 1996–2017 Biomedical Research Forum, LLC. All Rights Reserved.

Share this:
Facebooktwittergoogle_plusmailFacebooktwittergoogle_plusmail