Fig. 2.
Stability of 93F3 variants. (A and B) The melting curves for HALA, HGLG, HMLM, and HPLP are shown as measured by thermal scanning (solid circles) and light scattering [aggregation (agg); open circles]. The melting temperatures are fit directly to the melting curves in A or to Gaussian function fits to the derivative plots in B (Fig. S3 and SI Materials and Methods). (C) Both thermal scanning and light scattering show that the germ-line and mature antibodies have similar stabilities, but the antigen-binding mutations alone destabilize whereas the peripheral mutations alone stabilize the germ-line scaffold. HTTS, high-throughput thermal scanning. (D) Heat map of Fab Tms from the combination of heavy-chain and light-chain mutation groups. Note that much of the stabilization by the peripheral mutations comes from the H2 group. The high stabilities of L3 Fabs indicate that antigen-binding light-chain mutations N93D and N95P are responsible for most of the destabilization upon maturation.