The rate and equilibrium constants for a multistep reaction sequence for the aggregation of superoxide dismutase in amyotrophic lateral sclerosis

Sagar D. Khare, Michael Caplow, Nikolay V. Dokholyan

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Abstract

Mutation-induced aggregation of the dimeric enzyme Cu, Zn superoxide dismutase 1 (SOD1) has been implicated in the familial form of the disease amyotrophic lateral sclerosis, but the mechanism of aggregation is not known. Here, we show that in vitro SOD1 aggregation is a multistep reaction that minimally consists of dimer dissociation, metal loss from the monomers, and oligomerization of the apo-monomers: Dholokon koff2Mholokm-km+2M apokaggA where Dholo, Mholo, Mapo, and A are the holo-dimer, holo-monomer, apo-monomer, and aggregate, respectively. Under aggregation-promoting conditions (pH 3.5), the rate and equilibrium constants corresponding to each step are: (i) dimer dissociation, Kd ≈1 μM; koff ≈ 1 × 10-3 s-1, kon ≈1 × 103 M -1·s-1; (ii) metal loss, Km ≈ 0.1 μM, km- ≈ 1 × 10-3 s-1, km+ ≈ 1 × 104 M -1·s-1; and (iii) assembly (rate-limiting step), kagg ≈ 1 × 103 M-1·s -1. In contrast, under near-physiological conditions (pH 7.8), where aggregation is drastically reduced, dimer dissociation is less thermodynamically favorable: Kd ≈ 0.1 nM, and extremely slow: koff ≈ 3 × 10-5 s-1, kon ≈ 3 × 10 -1·s-1. Our results suggest that familial amyotrophic lateral sclerosis-linked SOD1 aggregation occurs by a mutation-induced increase in dimer dissociation and/or increase in apomonomer formation.

Original languageEnglish (US)
Pages (from-to)15094-15099
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume101
Issue number42
DOIs
StatePublished - Oct 19 2004

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