Hidden in the soil of backyards around the world, yet-to-be-discovered enzymes could be holding the key to improved gene therapies that could cure the most debilitating diseases plaguing modern society.

Thanks to the University of Delaware College of Engineering’s Kevin Solomon, it may be an undergraduate student who discovers the enzyme (or enzymes) that changes the world. But their search is just one part of a broad effort to pursue a new avenue for improving modern-day biomolecular engineering tools.

“The genomes of bacteria and other cells are simply a collection of genes (like words) made up of individual nucleotides (genetic alphabet),” Solomon said. “We want to develop technology that allows us to edit genomes or rewrite specific letters of that message to cure diseases or make economical biofuels without having to rewrite the whole page.”

Solomon, a synthetic biology expert and assistant professor in UD’s Department of Chemical and Biomolecular Engineering, was recently awarded about $1.1 million in funding for this work from the National Science Foundation’s Faculty Early Career Development (CAREER) program, one of the most prestigious awards a junior faculty member can receive.

“Bio-based processes in chemical engineering have been one of the biggest growth areas of the discipline in the last 30-40 years, and it’s so great to see young faculty like Kevin continuing to lead that charge,” said Department Chair Eric Furst. “At the University of Delaware, we pride ourselves in the access our students have to cutting-edge work in chemical engineering, where they can get involved with pushing that edge.”

The award will support five years of research and a new educational program, in partnership with Purdue University. By working with undergraduate students at these two universities, and potentially also with African citizens attending a unique graduate program at Purdue, the crowdsourced research element of this cross-collaborative effort could have international impact.

“This is a way to bring the excitement of research and development to students just starting down that path, who may someday become researchers and make groundbreaking discoveries themselves,” said Kari Clase, a professor of agricultural and biological engineering at Purdue who worked alongside Solomon before he joined the faculty at UD.

Solomon is an award-winning educator and researcher who joined UD in 2021 from Purdue University, where he was an assistant professor for five years. He received his master’s and doctoral degrees in chemical engineering from the Massachusetts Institute of Technology. In 2019, he was awarded the U.S. Department of Energy’s Early Career Award and also provided expert testimony before the U.S. House of Representatives on educational and federal funding priorities to advance the development of the bioeconomy.

“The research and development aspects he’s working on are likely to find their way into the solutions of tomorrow,” Clase said. “This prestigious award is also a recognition of the quality of his work and its potential impact.”

Bioengineering solutions

Solomon’s lab at UD is interested very broadly in sustainability and using biology to engineer solutions, Solomon said. They want to finetune the tools used in genetic engineering in order to produce a wide variety of biology-based improvements that could benefit society, from better medicines and treatments to more sustainable fuels to better, more nutritious foods.

The concept isn’t that different from how cheese or beer are made — it’s just that engineers like Solomon can use synthetic biology to reprogram cells to make them do whatever they want, such as create insulin, for example.

To do that reprogramming, they need “scissors,” or enzymes, that can cut DNA. An enzyme is simply a protein that can cause or accelerate chemical reactions or processes, such as metabolism, that occur in living organisms. Using these “scissors,” they can then paste anything they want in that space during the editing — so instead of a cell working to create beer, this engineered program tells it to make insulin or fuel or a more nutritious vegetable, Solomon explained.

The current scissors being used in the second generation of gene editing technology known as CRISPR are good, but they aren’t as precise as scientists would like them to be.

Think about it like a page of text, Solomon said. You have a tool that would allow you to edit any word in the document, as long as it’s placed next to the word “and.” That specificity is great, unless the cut that needs to be made is in the very first sentence at the top of the page — then you would have to cut and retype the entire page.

However, if the text is in Spanish or French instead of English, that same tool will not be effective at all because the words are different and are arranged differently. Think of it as if microbes speak many different languages, but right now we’re only good at engineering those that speak English.

Those limitations mean current tools cannot be successfully used on emerging organisms that have interesting or complex properties that may not be in the same “language” as the tools being used. Those different properties may also mean these organisms that cannot yet be genetically engineered may have the potential to solve the most complex problems society faces, from disease to nutrition to energy sources, Solomon said.

“We’re developing technologies that can make very clean and precise edits,” Solomon said. “While people are currently looking at engineering or finding new CRISPR-associated enzymes that recognize shorter or different ‘words,’ we’re looking at a different class of enzymes altogether.”

Introducing new enzymes

Prokaryotic Argonaute proteins (pAgos) are among a diverse group of enzymes that don’t rely on a specific DNA sequence motif or “word” like “and” for cutting, like the current enzymes used in CRISPR-based gene editing do, allowing for a level of precision not before seen in genetic engineering.

Returning to the analogy of text on a page, this new set of “scissors” can cut to edit any word on the page when programmed by a short DNA sequence, regardless of whether or not it’s anchored to another specific word, Solomon explained. That makes pAgos one type of understudied system that could help make the more precise cuts needed to work with more complex genomes, Solomon explained in his project proposal.

The challenge is that, while it has been established that this system works in theory, known pAgos generally require high temperatures. For example, the first well-studied variants of these enzymes operate at 60-80 degrees Celsius (140-158 degrees Fahrenheit), which would unfortunately kill most living cells that scientists want to engineer. There is evidence, however, that there are enzymes that can work in varying conditions (or more normal temperatures, like that of the human body) that might also work safely in gene therapy applications, Solomon said.

In 2021, Solomon and seven other researchers (six of whom were students in his lab) published their findings on another pAgo enzyme that requires salt, which will need significant development before use in humans, but did suggest there’s a real need to search for other existing possibilities.

This latest project is inspired by the idea of discovering those more promising examples and developing them as the “perfect molecular scissors,” Solomon said.

“Based on the similarity to those we’ve studied, we know there are a couple hundred organisms with these proteins across the globe, and at least 100 in conditions that wouldn’t kill us,” he said. “We want to find the ones that are the simplest to deploy, that cut very quickly and are accurate.”

Down in the dirt

The CAREER Award blends research and education, so Solomon also will be building new laboratory curricula and a pilot course in collaboration with Purdue University to support college sophomores hunting for these types of enzymes in the environment.

Researchers will pilot the undergraduate work within a course at Purdue that already exists and has about 70 enrolled students annually, Solomon said.

“The goal is to teach them how to do basic biotechnology while crowdsourcing the research with the potential to make new discoveries,” he said. Every student will work with soil samples, isolating and assembling different pieces of DNA that they will screen and analyze with technologies from the Solomon lab.

There are already programs and protocols that exist for this kind of work. At Purdue and other institutions linked with the Howard Hughes Medical Institute Science Education Alliance-Phage Hunters, for example, students already collect soil samples every year, isolating viruses that target bacteria. Work done by students at Purdue has collected novel datasets that can help inform strategies for treating increasingly antibiotic-resistant diseases like tuberculosis.

Instead of looking for random viruses, though, dozens of undergraduate biological engineering students will search for enzymes that could lead to better medicines, cheaper gasoline or better crops.

“We’re looking for needles in a haystack. But we have a good sieve,” Solomon said. “We’re handing sieves out to students to discover the next best pAgo or molecular scissors.”

Another graduate program in Clase’s department also works with African citizens from over 12 different countries who are professionals in manufacturing or regulatory positions who work to bring medical products to those in need. Solomon said he also hopes to engage these students in the project as well.

“As we move forward and develop this collaboration with Dr. Solomon there’s also the potential this work could be shared broadly with and benefit the research community,” Clase said. “The more we have a molecular understanding of things, the better we can understand how to alter it when it doesn’t go well. That has a huge, far-reaching potential.”