Super-compensator alleles rescue diverse deleterious substitutions and buffer the effects of mutations

Understanding loss-of-function mutations and their potential compensatory mechanisms is crucial for elucidating the complexities of gene expression. Here, we investigate whether additional mutations can rescue non-functional alleles and their evolutionary significance. To achieve this, we conducted an extensive analysis of 372,867 amino acid sequence variants of the imidazole glycerol-phosphate dehydratase (IGPD, PDB: 6ezm) protein, which is over 1000 times greater than previously examined. We discovered that one or more additional substitutions located in loop and interface regions could easily restore function for each of the 10,019 of 31,701 (31.6%) non-functional missense allele. We further identified a small set of super-compensatory substitutions that can increase the function of many diverse alleles and flatten the fitness landscape, that is, buffer the fitness effects of both beneficial and deleterious substitutions. We also found that the protein structures of rescuable variants are more stable than those of non-rescuable variants. This suggests that restoring the stability of protein structures is a key part of compensation. These findings offer new insights into the understanding of protein function loss and its adaptive compensatory evolution.