Melissa P Tumen-Velasquez, Christopher W Johnson, A. Ahmed, Graham Dominick, E. Fulk, P. Khanna, Sarah A. Lee, A. L. Schmidt, Jeffrey G. Linger, M. Eiteman, G. Beckham, E. Neidle
Proceedings of the National Academy of Sciences, 2018
Significance Gene duplication and divergence are cornerstones of evolution. Genetic redundancy resulting from repeated DNA provides flexibility for transient changes in copy number that may confer selective benefit under new or changing environmental conditions. This work describes a method for creating tandem arrays of specific DNA segments in a bacterium, Acinetobacter baylyi, to accelerate experimental evolution. The induced chromosomal gene amplification mimics a natural process that would otherwise occur more slowly and stochastically. The success of this approach for the evolution of novel protein function was demonstrated with studies of an enzyme that has bioenergy applications in lignin valorization. A protein variant, which emerged from Evolution by Amplification and Synthetic biology, is beneficial in a different bacterium, indicating the broad utility of the method. Experimental evolution is a critical tool in many disciplines, including metabolic engineering and synthetic biology. However, current methods rely on the chance occurrence of a key step that can dramatically accelerate evolution in natural systems, namely increased gene dosage. Our studies sought to induce the targeted amplification of chromosomal segments to facilitate rapid evolution. Since increased gene dosage confers novel phenotypes and genetic redundancy, we developed a method, Evolution by Amplification and Synthetic Biology (EASy), to create tandem arrays of chromosomal regions. In Acinetobacter baylyi, EASy was demonstrated on an important bioenergy problem, the catabolism of lignin-derived aromatic compounds. The initial focus on guaiacol (2-methoxyphenol), a common lignin degradation product, led to the discovery of Amycolatopsis genes (gcoAB) encoding a cytochrome P450 enzyme that converts guaiacol to catechol. However, chromosomal integration of gcoAB in Pseudomonas putida or A. baylyi did not enable guaiacol to be used as the sole carbon source despite catechol being a growth substrate. In ∼1,000 generations, EASy yielded alleles that in single chromosomal copy confer growth on guaiacol. Different variants emerged, including fusions between GcoA and CatA (catechol 1,2-dioxygenase). This study illustrates the power of harnessing chromosomal gene amplification to accelerate the evolution of desirable traits.
Cited by 20 publications.
Field of study: Biology