Looking for an enzyme in a haystack

Dr. Kristala Prather describes the societal imperative prompting her research succinctly: “We have to stop digging up dead dinosaurs to make the world run. In other words, we have to get away from using fossil fuels and fossil inputs to produce the molecules we need for everyday life.” How, then, are we to synthesize molecules that have the right properties for their intended use? According to Dr. Prather, biology is a master chemist, capable of generating a diverse array of useful molecules. Her research proposes to refine and expand this capability by helping nature to evolve biosensors able to analyze a large amount of enzymes and identify the ones with the most relevant characteristics.

Finding just the right enzymes is a significant challenge, however. A method of altering enzyme traits known as directed evolution (for which Caltech’s Frances Arnold won the 2018 Nobel Prize in Chemistry) requires extremely large libraries of DNA sequences to be generated and screened—connecting the size of these libraries with sufficient throughput for screening is a major barrier. Two screening methods could be practical: one would utilize some kind of biosensor, the core element of which would be a transcription factor that responds favorably to the metabolite of interest and produces an easily detectable signal, such as fluorescence; the other would use the transcription factor to control expression of an essential gene, with best candidates identified from a standard enrichment culture. Dr. Prather’s novel approach to access such sensors is to expose half a dozen types of bacteria to half a dozen unique molecules, and through this exposure allow microbial cells to develop new sensors on their own; she will observe this action by measuring RNA levels and changes in gene expression.

Speeding along evolution

“My central premise—that nature will both enhance and, if necessary, evolve metabolite-protein interactions through prolonged exposure—is a high-risk one,” says Dr. Prather. “After all, evolution occurs over very long time scales and there is no preliminary data to support my hypothesis. That’s why this work would almost certainly not be funded by traditional sources.” There are also technical challenges involved in this work, such as determining the right level of exposure and maintaining a sterile environment. In addition, a lot of inference is involved in deciding whether or not observed changes are actually connected to something useful. However, it has been proven that prolonged exposure of a microbial culture to a target molecule can result in the evolution of a relevant sensor (in Japan in 2016, a bacterium was discovered that had naturally evolved to consume plastic).

Uncommon sensors for the common good

The findings from this work could have broader impacts beyond those mentioned above. For example, the search for effective enzymes need not be limited to a goal of creating biological synthesis. Using these methods, microbial systems could be engineered to enhance the degradation of both toxic and non-toxic wastes (as in the Japan example above). Likewise, the approach for sensor development could be used for detecting small molecules in the environment or to be incorporated into microbial diagnostics. Sensors can further be utilized in the development of complex microbial systems that respond to varying environmental conditions by activating different cellular responses.