Researchers from Michigan State University in East Lansing have created a synthetic nano-sized factory based on natural ones found in bacteria. The factories help the bacteria in different ways – some make nutrients, and others sequester toxic materials that would otherwise make the bacteria sick.
The researchers say the synthetic factories may one day be used in medical, industrial, or bioenergy applications.
While the factories serve different purposes, they all have shells made of protein tiles. The researchers wanted to be able to direct useful enzymes to the factories by attaching them to the tail ends of the proteins that make up the shells.
However, the ends, or termini, of most shell protein tiles face the outside of the factories, so molecules fused to the protein ends remained on the outside surface instead of the inside. This is a problem if the goal is to keep enzymes inside a factory separate from the rest of the cell.
“In order to send proteins to the inside of the factory, we needed a new kind of building block that still assembled into shells,” says Bryan Ferlez, a post doctorate researcher in the Kerfeld lab. “We aimed to redesign a shell protein so its termini face the inside. The end result is that cargo connected to this shell protein would also end up inside the shell.”
In the study, the scientists took the most abundant shell protein, BMC-H, and turned it inside out through a technique called circular permutation.
The segments of the amino acid sequence were shuffled and the ends were glued together. The new termini were introduced on the inner face of the protein, resulting in a new, synthetic shell protein that looks almost identical to its natural counterpart but with ends facing the inside.
The new structure is a building block to construct factory shells. The scientists have produced factory shells with the new protein that are similar in size and appearance to the originals. The new structure can incorporate molecules inside the shell. The team tested the concept by fusing a fluorescent cargo protein to the new BMC-H protein. Microscopy and biochemical testing show the cargo inside the shell.
“By making more or less of the new BMC-H protein with a fluorescent protein fused to its terminus, we were also able to control the amount of cargo that incorporates into the shell,” says Ferlez.
Next, Ferlez wants to target useful molecules into a synthetic factory made with the new shell protein.
“We can start to build metabolic pathways, or assembly lines, and define the amounts and locations of enzymes within these nanofactories,” he says. “Someday, we could use this system to enhance the production of rubber, biofuels, and other commodities.”
The work was published in the journal Metabolic Engineering.