Increasing and decreasing protein stability: Effects of revertant substitutions on the thermal denaturation of phage λ repressor

The thermal denaturations of five revertant λ repressors containing single amino acid substitutions in their N‐terminal domains have been studied by differential scanning calorimetry. Two substitutions slightly decrease stability, and the remaining three render the protein more stable than wild type. The Gly₄₈ → Asn and Gly₄₈ → Ser proteins are 4°C more stable than wild type. These two substitutions replace an α helical residue, and in each case a poor helix forming residue, glycine, is replaced by a residue with a higher helical propensity. We also present data showing that one revertant, Tyr₂₂ → Phe, has reduced operator DNA binding affinity despite its enhanced stability.