Cell-Free Systems Accelerate Protein Engineering and Biosensor Development

Michael Jewett, a bioengineer at Stanford University, has long grappled with the inherent conflicts of using synthetic biology for research. While he relies on microorganisms to manufacture specific compounds or follow genetic instructions, these evolved organisms often prioritize their own survival over experimental goals, sometimes deactivating foreign DNA or diverting resources.

To resolve this, Jewett and other researchers have turned to cell-free protein expression systems. Rather than fighting the ‘tug of war’ with living cells, they strip away cell walls and utilize the internal molecular machinery as a standalone factory. Although this approach traces its roots to the 1960s, when it helped decipher the genetic code, it is experiencing a resurgence. Vincent Noireaux, a biophysicist at the University of Minnesota, describes it as a current ‘in’ trend, noting a proliferation of new applications and properties.

These systems, which can be derived from bacterial or eukaryotic lysates or assembled from purified components known as Protein Synthesis Using Recombinant Elements (PURE), offer significant speed advantages. Wilson Wong, a synthetic biologist at Boston University, explains that the design-build-test-learn cycle is compressed from days or weeks into hours. This allows scientists to screen thousands of protein options and produce substances that would be toxic to or altered by living cells.

However, cost and scalability remain hurdles. Running a liter-scale cell-free system can exceed $4,000 in reagents, making it roughly ten times more expensive than cell-based methods. Additionally, because these systems exhaust their energy supplies quickly, reactions must typically conclude within a few hours. To mitigate costs, some researchers operate at sub-microlitre scales, while others, like Jewett’s team, have optimized formulations to reduce expenses by 95%, bringing the cost to under $100 per gram of protein produced.

Despite these challenges, cell-free systems excel in rapid biosensor development. Jewett and his collaborator Julius Lucks from Northwestern University have developed sensitive detectors for environmental contaminants. In 2020, they introduced ROSALIND, a system that uses DNA-binding transcription factors to detect molecules like copper and zinc. More recently, they utilized machine learning to refine a sensor for lead, achieving detection limits of 5.7 parts per billion after just a few iterative rounds of design and testing in cell-free environments.

The commercial landscape is also expanding. Companies such as New England Biolabs, Daicel Arbor Biosciences, Ginkgo Bioworks, and GenScript offer various cell-free kits. Emily Chen of New England Biolabs notes that lysate-based systems provide high yields at lower costs, making them accessible for startups. Meanwhile, researchers like Henrike Niederholtmeyer at the Technical University of Munich are exploring plant-based systems. By using chloroplast lysates from spinach and other plants, scientists can test transgenes for chloroplast engineering without waiting for slow-growing plants, potentially streamlining the development of genetically modified crops.