α-Synuclein aggregation variable temperature and variable pH kinetic data: A re-analysis using the Finke–Watzky 2-step model of nucleation and autocatalytic growth Original Research Article

The aggregation of proteins is believed to be intimately connected to many neurodegenerative disorders. We recently reported an “Ockhamʹs razor”/minimalistic approach to analyze the kinetic data of protein aggregation using the Finke–Watzky (F–W) 2-step model of nucleation (A → B, rate constant k1) and autocatalytic growth (A + B → 2B, rate constant k2). With that kinetic model we have analyzed 41 representative protein aggregation data sets in two recent publications, including amyloid β, α-synuclein, polyglutamine, and prion proteins (Morris, A. M., et al. (2008) Biochemistry 47, 2413-2427; Watzky, M. A., et al. (2008) Biochemistry 47, 10790–10800). Herein we use the F–W model to reanalyze protein aggregation kinetic data obtained under the experimental conditions of variable temperature or pH 2.0 to 8.5. We provide the average nucleation (k1) and growth (k2) rate constants and correlations with variable temperature or varying pH for the protein α-synuclein. From the variable temperature data, activation parameters ΔG‡, ΔH‡, and ΔS‡ are provided for nucleation and growth, and those values are compared to the available parameters reported in the previous literature determined using an empirical method. Our activation parameters suggest that nucleation and growth are energetically similar for α-synuclein aggregation (ΔG‡nucleation = 23(3) kcal/mol; ΔG‡growth = 22(1) kcal/mol at 37 °C). From the variable pH data, the F–W analyses show a maximal k1 value at pH ~ 3, as well as minimal k1 near the isoelectric point (pI) of α-synuclein. Since solubility and net charge are minimized at the pI, either or both of these factors may be important in determining the kinetics of the nucleation step. On the other hand, the k2 values increase with decreasing pH (i.e., do not appear to have a minimum or maximum near the pI) which, when combined with the k1 vs. pH (and pI) data, suggest that solubility and charge are less important factors for growth, and that charge is important in the k1, nucleation step of α-synuclein. The chemically well-defined nucleation (k1) rate constants obtained from the F–W analysis are, as expected, different than the 1/lag-time empirical constants previously obtained. However, k2 × [A]0 (where k2 is the rate constant for autocatalytic growth and [A]0 is the initial protein concentration) is related to the empirical constant, kapp obtained previously. Overall, the average nucleation and average growth rate constants for α-synuclein aggregation as a function of pH and variable temperature have been quantitated. Those values support the previously suggested formation of a partially folded intermediate that promotes aggregation under high temperature or acidic conditions.