Calorimetric Analysis of Thermodynamic Stability and Aggregation for Apo and Holo Amyotrophic Lateral Sclerosis-associated Gly-93 Mutants of Superoxide Dismutase *

摘要: Differential scanning calorimetry was used to measure changes in thermodynamic stability and aggregation for glycine 93 mutants of human copper, zinc-superoxide dismutase (SOD). Glycine is a conserved residue at position i + 3 tight turn has been found be mutational hot spot familial amyotrophic lateral sclerosis (fALS). The fALS-associated mutations, G93A, G93S, G93R, G93D, G93V, were made pseudo wild-type background containing no free cysteines, which prevented the formation aberrant disulfide bonds upon thermal unfolding, enabled quantitative analysis effects mutations. Thermal unfolding highly reversible all SODs both fully metallated (holo) metal-free (apo) forms. data holo-SODs apo-pseudo-wild-type SOD well fit by 2-state model native dimer (N2) two unfolded monomers (2U), N2 ↔ 2U. holo- apo-forms are significantly destabilized (by 1.5–3.5 kcal mol–1 monomer) relative corresponding forms wild-type, with stabilities being correlated statistical preferences amino acids this structural context. Although van't Hoff (ΔHvH) calorimetric (ΔHcal) enthalpy ratios close unity apo-pseudo-wild-type, consistent transition, ΔHvH considerably larger than ΔHcal apo-mutants. This suggests that mutations cause apo-SOD have an increased propensity misfold or aggregate, may linked toxic mutant fALS.