Ancestral sequence reconstruction relies on phylogeny and statistics to infer the most likely amino acid
Scientists collect sequences from databanks of the modern versions of the protein of interest from different organisms.
Computer algorithms construct a phylogenetic tree for the proteins (Curr Opin Struct Biol, 38:37–43, 2016).
The programs can then infer the sequences that likely existed at nodes of the tree, before the modern species evolved.
Finally, the scientists order synthetic DNA and generate those proteins in the lab to use for experiments.
One way to ensure that an ASR protein behaves like the true ancestor is to resurrect and test not only the best amino acid sequence generated by the algorithms, but a few proteins with the second-best guesses, or third-best guesses, and so on. If those alternative ancestors act like the best-guess version, then researchers figure the conclusions are probably robust. Recently, evolutionary synthetic biologist Eric Gaucher of Georgia State University tested ASR accuracy in a different way. He generated an entirely artificial phylogenetic tree, starting with red fluorescent protein and randomly mutating it to evolve 19 diversely colored fluorescent proteins. Then he used ASR to predict the ancestor of those 19 descendants, and compared the results to the true ancestors. The results were reassuring. Overall, the five different ASR algorithms he tried identified the ancestral sequence with about 97 percent accuracy (Nat Commun, 5:12847, 2016).
The Perfect Starting Point
Bioengineers love resurrected proteins because they often combine two desirable features: thermostability and promiscuity. For example, researchers at the University of Granada in Spain reconstructed several versions of an antibiotic-resistance protein called beta-lactamase, going back as far as 3 million years. As the protein evolved, its melting point dropped from more than 80 °C to less than 60 °C. It also became more specific for penicillin, losing its ability to neutralize other drugs (J Am Chem Soc, 135:2899–902, 2013).
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