Figure 1.

Subunit exchange measures the stability of recombinant TTR tetramers. (A) The top left panel shows a ribbon diagram of the TTR tetramer with individual subunits shown in different colors. Arrowheads indicate the thyroxine-binding pockets. The top right panel shows a cartoon of the same tetramer. The weak dimer–dimer interface is indicated with a dotted line. The bottom panel shows a schematic of the steps involved in dissociation, reassociation, and subunit exchange of untagged WT TTR subunits (green squares) and dual-FLAG-tagged (FT₂) WT TTR subunits (orange squares). (B) Recombinant WT and FT₂·WT TTR were mixed in equal amounts and separated by ion exchange chromatography after exchange for 7 days. The initial populations of homotetramers mix to create heterotetramers incorporating zero to four FT₂·WT TTR subunits. Green squares represent untagged WT TTR subunits and orange squares FT₂·WT TTR subunits. (C) To quantify subunit exchange, the relative area of peak 1 at time zero is calculated and used in the binomial equation to calculate the expected area of peak 3 at equilibrium. At each time point, the fraction exchange is calculated by comparing the actual relative area of peak 3 at that time point to the expected relative area of peak 3 at equilibrium (see the text for details).