Evolutionary Engineering a Larger Porin Using a Loop-to-Hairpin Mechanism

In protein evolution, diversification is generally driven by genetic duplication. The hallmarks of this mech-anism are visible in the repeating topology of various proteins. In outer membrane (3-barrels, duplication is visible with (3-hairpins as the repeating unit of the barrel. In contrast to the overall use of duplication in diversification, a computational study hypothesized evolutionary mechanisms other than hairpin duplica-tions leading to increases in the number of strands in outer membrane (3-barrels. Specifically, the topology of some 16-and 18-stranded (3-barrels appear to have evolved through a loop to (3-hairpin transition. Here we test this novel evolutionary mechanism by creating a chimeric protein from an 18-stranded (3-barrel and an evolutionarily related 16-stranded (3-barrel. The chimeric combination of the two was created by replacing loop L3 of the 16-stranded barrel with the sequentially matched transmembrane (3-hairpin region of the 18-stranded barrel. We find the resulting chimeric protein is stable and has characteristics of increased strand number. This study provides the first experimental evidence supporting the evolution through a loop to (3-hairpin transition.(c) 2023 The Author(s). Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://crea-tivecommons.org/licenses/by-nc-nd/4.0/).

Automated summary

In protein evolution, diversification is often driven by genetic duplication. However, a computational study suggests that another evolutionary mechanism involving a transition from loops to hairpins can lead to an increase in the number of strands in outer membrane proteins.