The consequences of cavity creation on the folding landscape of a repeat protein depend upon context
Extant protein sequences are the result of evolutionary pressure. Eliminating core packing interactions by mutation, regardless of position, generally results in similar perturbations to global stability. In contrast, we find that cavity creation has highly varied consequences for a protein folding landscape, depending upon the context in which the cavities are introduced. These observations have implications for interpreting evolutionary adaptation, as it is likely that proteins have evolved to exhibit optimal levels of conformational heterogeneity and dynamics. These results should also inform protein engineering efforts, as they provide insight into how sequence can modulate the population of functionally important excited states, as well as states that lead to secondary, undesirable reactions such as oligomerization, aggregation, surface activity, and phase separation.