Proc. Natl. Acad. Sci. USA, published online 16 December 2011, doi:10.1073/pnas.1111537108

Credit: ANDERS SANDSTRÖM

Enzymes have been used by humans to carry out chemical transformations on small-molecule substrates for hundreds of years—for example, for brewing beer or making cheese. However, the high substrate specificity of many enzymes makes it challenging to generate new 'biocatalysts' that efficiently carry out chemical transformations on non-native substrates. Sandström et al. now show that it is possible to generate a very small library of mutant lipases and still find a highly active variant that is able to perform an enantioselective reaction on a larger non-native substrate. The authors selected nine amino acids in the substrate-binding pocket for simultaneous mutation, making a library that only contained 1,024 variants. The most active variant identified had mutations at five of the targeted positions. An analysis of single and double mutants suggested that the native enzyme lay in a relatively 'flat' or inactive region of the protein fitness landscape, and the authors believe that 'walking' (via directed evolution) from that position would not have yielded a highly enantioselective mutant lipase for the new substrate. However, that several mutations were simultaneously incorporated meant that the authors had 'hopped' to a different, more active region of the protein fitness landscape. This approach may facilitate the discovery of new biocatalysts for other enantioselective chemical transformations.