The impact of amino acid insertions and deletions on protein stability and function

Amino acid insertions and deletions (indels) are common mutational events but there is limited information about their effects on protein stability and function. Whereas substitutions alter amino acid side chains, indels are ‘backbone mutations’ that modify protein length with potentially severe and difficult-to-predict outcomes. To address the sparsity of data, we quantified indel effects on the in vivo abundance of diverse protein domains using a deep-sequencing- based protein fragment complementation assay. We use this data, together with an existing large-scale indel mutagenesis dataset on in vitro protein stability, to examine indel tolerance across protein domains and secondary structures, comparing how patterns of indel tolerance differ from those of substitutions. Additionally, our analysis shows that current computational methods poorly predict indel effects. To address this, we present INDELi, a series of models that integrate experimental or predicted substitution effects with secondary structure information to provide better predictions of indel effects on both protein stability and pathogenicity. Lastly, we explore the impact of indels on protein-protein interactions, showing that while frequently detrimental to protein binding, 1aa insertions can also act as an important class of gain-of-function variants. Using abundance and binding data, we further quantify allosteric communication across three protein-protein interactions, revealing that insertions and deletions can have long-range and allosteric effects on binding affinity. In summary, our results provide insights into the impact of indels on protein stability and function, as well as a method to predict indel tolerance genome-wide.