Cell-free biotechnology is rapidly emerging as a powerful production platform that can be used to manufacture industrial enzymes, therapeutics, anti-microbial peptides, industrial chemicals and biomaterials. Cell-free biotechnology is currently dominated by systems that are based on extracts produced from cells that have been mechanically lysed to smash them open, thus extracting (freeing) their contents. Essentially, cellular extracts include the metabolic enzymes that cells use to transform sugars into cellular energy (e.g. ATP) and the machinery (e.g. polymerases and ribosomes) that cells use to produce proteins according to the instructions found within its DNA. The freeing of this cellular machinery, or “gloop” as Hal Hodson calls it in his recent article “Cell-free biotech will make for better products” is potentially transformative for industrial biotechnology, since the extracted cellular machinery does not have to waste resources on unproductive activities (e.g. cell growth) and can instead be wholly directed towards the production of useful products. Equally, removing the cell membrane means that cell-free biotechnologists have unparalleled access to the cellular machinery and therefore, programming cell-free systems to produce specific proteins, natural products, biomaterials and chemicals is as simple as adding some energy components (e.g. sugars and amino acids) and some engineered DNA to the cell extract.

Recent advances in synthetic biology, have made it relatively easy to produce massive libraries of engineered DNAs that might encode different versions of the instructions needed for cells (or a cell extract) to make, say a therapeutic antibody or bioplastics. Indeed, SynbiCITE’s own London DNA Foundry can readily assemble thousands of new engineered DNAs and LabGenius, a SynbiCITE supported start-up has proprietary technologies that can produce engineered DNA libraries that consist of up to 10¹³ unique variants. But, testing these vast libraries of engineered DNA variants to find the best designs can be slow and cumbersome using cells. It can sometimes take days to get an engineered DNA into a cell and even if you succeed the cells may try to ‘turn-off’ the burdensome engineered DNA so that they can continue to use their cellular resources for growth. These limitations are circumvented in cell-free systems that are readily able to execute many different engineered DNA programmes, even those that are highly toxic to living bacterial cells (e.g. anti-microbial peptides). Whilst large scale cell-free biomanufacturing reactions are possible, smaller scale cell-free reactions can also be incredibly powerful. For instance, thousands of extremely small volume cell-free reactions can be run in parallel, making cell-free the perfect low-cost engineered DNA screening platform. This means that cell-free systems have the potential to accelerate biotechnological discovery making it faster and cheaper to discover new therapeutics, industrial chemicals and other bio-products (e.g. biomaterials).

Whilst cell-free systems may be the superior choice for several applications, cell-free systems can also complement cell-based biomanufacturing efforts (e.g. microbial fermentation). Cell-free research in our laboratory at Imperial College, has been influential in demonstrating comparability between the activities of engineered DNA designs tested in cells and cell-free systems. In other words, cell-free extracts retain many of the biochemical characteristics of the cells from which they were derived and thus cell-free experiments can inform how the engineered DNA may function in the original cell. Indeed, the Fremont lab has demonstrated in vitro (cell-free) and in vivo (cell) comparability of engineered DNA designs in several different bacterial cells and their derivative cell-free systems (including Escherichia coli and Bacillus subtilis). Moving forward the Freemont group is bioprospecting (testing) the potential of cell extracts, from diverse organisms that have unique biochemistries. The goal is to engineer novel and highly robust cell-free systems that can be used as platforms for medical biosensors in global health or in another ongoing project to produce biomaterials (e.g. PHA based bioplastics).

Innovations in cell-free technologies are also emerging from new entrants. Established cell-free biotechnology companies, including Sutro Biopharma and GreenLight Biosciences are increasingly facing competition from the synthetic biology community in this rapidly developing field. Indeed, several SynbiCITE supported researchers, pre-startups and startups are carrying out proof-of-concept projects to develop innovative cell-free technologies that have the potential to disrupt incumbent companies in many different industries – ranging from medical diagnostics and therapeutics, to biomaterials. It’s now time for biotechnology to free itself of the cell.

Authors: Richard Kelwick and Paul Freemont