title: Combined Isothermal Titration and Differential Scanning Calorimetry Define Three-State Thermodynamics of fALS-Associated Mutant Apo SOD1 Dimers and an Increased Population of Folded Monomer.
abstract: Many proteins are naturally homooligomers, homodimers most frequently. The overall stability of oligomeric proteins may be described in terms of the stability of the constituent monomers and the stability of their association; together, these stabilities determine the populations of different monomer and associated species, which generally have different roles in the function or dysfunction of the protein. Here we show how a new combined calorimetry approach, using isothermal titration calorimetry to define monomer association energetics together with differential scanning calorimetry to measure total energetics of oligomer unfolding, can be used to analyze homodimeric unmetalated (apo) superoxide dismutase (SOD1) and determine the effects on the stability of structurally diverse mutations associated with amyotrophic lateral sclerosis (ALS). Despite being located throughout the protein, all mutations studied weaken the dimer interface, while concomitantly either decreasing or increasing the marginal stability of the monomer. Analysis of the populations of dimer, monomer, and unfolded monomer under physiological conditions of temperature, pH, and protein concentration shows that all mutations promote the formation of folded monomers. These findings may help rationalize the key roles proposed for monomer forms of SOD1 in neurotoxic aggregation in ALS, as well as roles for other forms of SOD1. Thus, the results obtained here provide a valuable approach for the quantitative analysis of homooligomeric protein stabilities, which can be used to elucidate the natural and aberrant roles of different forms of these proteins and to improve methods for predicting protein stabilities.
authors.affiliation: Department of Chemistry, University of Waterloo , Waterloo, Ontario N2L 3G1, Canada.
authors.forename: Helen R
authors.initials: HR
authors.lastname: Broom
authors.affiliation: Department of Chemistry, University of Waterloo , Waterloo, Ontario N2L 3G1, Canada.
authors.forename: Kenrick A
authors.initials: KA
authors.lastname: Vassall
authors.affiliation: Department of Chemistry, University of Waterloo , Waterloo, Ontario N2L 3G1, Canada.
authors.forename: Jessica A O
authors.initials: JA
authors.lastname: Rumfeldt
authors.affiliation: Department of Chemistry, University of Waterloo , Waterloo, Ontario N2L 3G1, Canada.
authors.forename: Colleen M
authors.initials: CM
authors.lastname: Doyle
authors.affiliation: Department of Chemistry, University of Waterloo , Waterloo, Ontario N2L 3G1, Canada.
authors.forename: Ming Sze
authors.initials: MS
authors.lastname: Tong
authors.affiliation: Department of Chemistry, University of Waterloo , Waterloo, Ontario N2L 3G1, Canada.
authors.forename: Julia M
authors.initials: JM
authors.lastname: Bonner
authors.affiliation: Department of Chemistry, University of Waterloo , Waterloo, Ontario N2L 3G1, Canada.
authors.forename: Elizabeth M
authors.initials: EM
authors.lastname: Meiering
mesh_terms: D000690:Amyotrophic Lateral Sclerosis / Q000235:genetics*; D001051:Apoenzymes / Q000737:chemistry / Q000235:genetics; D002151:Calorimetry / Q000379:methods; D004795:Enzyme Stability; D006801:Humans; D009154:Mutation; D017510:Protein Folding; D055503:Protein Multimerization; D013482:Superoxide Dismutase / Q000737:chemistry* / Q000235:genetics; D000072105:Superoxide Dismutase-1; D013816:Thermodynamics
chemical_list: D001051:Apoenzymes; C000606290:SOD1 protein, human; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1