Impact of ancestral sequence reconstruction on mechanistic and structural enzymology

Ancestral sequence reconstruction (ASR) provides insight into the changes within a protein sequence across evolution. More specifically, it can illustrate how specific amino acid changes give rise to different phenotypes within a protein family. Over the last few decades it has established itself as a powerful technique for revealing molecular common denominators that govern enzyme function. Here, we describe the strength of ASR in unveiling catalytic mechanisms and emerging phenotypes for a range of different proteins, also highlighting biotechnological applications the methodology can provide. [Display omitted] •Ancestral Sequence Reconstruction is becoming a widely used tool to dissect the mechanisms of enzyme catalysis.•Historical analysis provides valuable data for biotechnology and biomedicine enzyme application.•A small number of substitutions are sufficient to generate or specialise enzyme functions.•Long distance substitutions play a key role in defining enzyme functionality.