At Northwestern University’s Jewett Lab in the Center for Synthetic Biology, researchers aim to create sustainable chemicals and materials out of existing organic compounds. Using cell-free metabolic engineering, they isolate the structural components from existing organisms, such as E. coli, and manipulate them to create new compounds. These types of reactions are called “cell-free” because they occur outside the confines of a cell.
“We focus on E. coli because it is super well-studied,” said Ashty Karim, research fellow and assistant scientific director at the Jewett Lab. “We know a lot about how it functions and how to manipulate it to do our engineering objectives.”
The lab’s engineering objectives are to create sustainable and renewable chemicals that can be used for biofuels and in manufacturing.
Ashty Karim, research fellow and assistant scientific director at the Jewett Lab. Karim develops cell-free systems for prototyping biosynthetic pathways to discover natural products such as sustainable biofuels. (Valerie Nikolas/MEDILL REPORTS) The lab’s back room is filled with freezers set to -80 degrees Celsius. This is where the researchers keep cryogenically frozen bacteria samples. (Valerie Nikolas/MEDILL REPORTS) Ashty Karim takes a sample of E. coli from one of the freezers in the back room. Before placing samples in the freezers, researchers flash freeze them with liquid nitrogen, which cryogenically preserves the bacteria and prevents them from dying at such low temperatures. (Valerie Nikolas/MEDILL REPORTS) Ashty Karim holds up a plate used with the lab’s Echo Liquid Handler. The Echo is a robotic machine that pipettes liquid chemicals and lysates. It uses acoustic waves to move liquids from one plate, such as the one Karim is holding, to another plate. By sending energy waves through the liquid, the Echo machine makes the liquids move, form droplets, and then transfer from one plate to another. This form of pipetting is not only more precise and efficient than doing it by hand, it also eliminates waste from plastic pipettes. (Valerie Nikolas/MEDILL REPORTS) Karim (right) with postdoctoral associate Markus Jeschek making E. coli extracts. After centrifuging an E. coli culture down to form a pellet and removing all of the liquid media, they transfer a wash buffer containing salts onto the E. coli cell pellets. After that they resuspend and mix the solution to wash the cells, getting them ready to be broken open to make extracts. (Valerie Nikolas/MEDILL REPORTS) Postdoctoral associate Maria Cabezas began working at the Jewett Lab in July 2018. Cabezas uses high-throughput technologies for engineering biology using cell-free systems. (Valerie Nikolas/MEDILL REPORTS) Maria Cabezas sets up reactions for a cell-free protein system in a Vibrio bacteria strain. (Valerie Nikolas/MEDILL REPORTS) Postdoctoral associate Bastian Vögeli joined the Jewett Lab just two months ago, after arriving from Switzerland. Vögeli’s research focuses on creating compounds such as decanol that can be used as alternative fuel sources. “We take E. coli, we lyse them – basically just take their guts, their innards – and let them make different single enzymes,” Vögeli says. Then he feeds sugars to the enzymes to make different types of fatty acids that serve as the building blocks for fuel. (Valerie Nikolas/MEDILL REPORTS) Vögeli adding a sample to the High Performance Liquid Chromatography (HPLC) machine, which researchers use to determine which molecules are present in mixtures. “Depending on what molecules are in there, it changes how the light passes through,” says Vögeli. The HPLC separates compounds by chromatography. “Basically, it’s a glorified pump,” Vögeli says. The HPLC mixes chemicals with solvents, such as water, methanol or sulfuric acid. As the samples cycle through the machine they separate and the HPLC gives a measure of how much light is shining through the liquid. Measuring the amount of light can help researchers determine which molecules are present in the sample. (Valerie Nikolas/MEDILL REPORTS) Researchers also take measurements using the gas chromatographer. The wire pictured is a 30-meter column that’s as thin as a capillary. As molecules travel through the column they are separated, thus giving researchers a better indication of which molecules are present in a mixture. (Valerie Nikolas/MEDILL REPORTS) Lab manager Lauren Clark isolates plasma DNA from E. coli. Clark has worked in the Jewett lab for more than five years. (Valerie Nikolas/MEDILL REPORTS) Clark waters the tobacco plants she uses to study chloroplasts. Tobacco plants are optimal for this type of research since they yield lots of leaves. “My samples are very much reduced in volume,” Clark says. “For every 300 grams of leaves, I might get three grams of chloroplasts.” To isolate the chloroplasts, Lauren rips open the leaves and puts them in a blender, which frees the chloroplasts. She strains the paste and then places it through a centrifuge, which separates out intact chloroplasts. The result produces cell free chloroplasts that Clark can use to study. (Valerie Nikolas/MEDILL REPORTS) Graduate student Blake Rasor uses cell-free metabolic engineering for biosynthesis. (Valerie Nikolas/MEDILL REPORTS) Blake Rasor sets up reactions to make acetone and butanediol from cell free lysates. Acetone and butanediol are two types of “commodity chemicals,” or compounds that can be used for making fuel or as additives. Commodity chemicals can be derivatized into other compounds as well. (Valerie Nikolas/MEDILL REPORTS)
Ashty Karim, research fellow and assistant scientific director at the Jewett Lab, holds up a sample of cryogenically frozen E. coli. (Valerie Nikolas/MEDILL REPORTS)