Proteins In Small Packages

Someday doctors may turn on tiny factories to churn out proteins inside patients by flicking a light switch. Researchers have stuffed lipid vesicles with the biochemical components for protein synthesis, and turned on production with light (Nano Lett., DOI: 10.1021/nl2036047). They hope the particles could deliver the drugs on demand to the parts of the body where they’re needed most.

To avoid side effects, scientists and doctors have long wanted to send a drug only to the tissues that need it, says Daniel Anderson of the Massachusetts Institute of Technology. He wondered if he could build lipid vesicles that wouldn’t just deliver but could actually make a drug “only where and when needed,” he says.

The first step was to get vesicles to make protein. The ingredients for protein synthesis came ready-made from a bacterial extract containing enzymes, ribosomes, nucleotides, amino acids, tRNA, and other materials. Another essential ingredient was a string of DNA containing a gene that encodes the desired protein.

The researchers combined a plasmid containing the gene for green florescent protein with the bacterial extract, and then added a phospholipid, which spontaneously formed vesicles by encapsulating the DNA and the extract. They selected a phospholipid that creates soft bilayers, so that protein can eventually escape from the particles. The researchers used confocal microscopy to observe the vesicles and within minutes, the particles began to glow bright green, indicating that they were making properly folded protein.

To gain control over the onset of protein synthesis, the researchers turned to a previously developed technique to trigger protein production with light (J. Biol. Chem., DOI: 10.1074/jbc.274.30.20895). They attached a so-called cage molecule, 1-(4,5-dimethoxy-2-nitrophenyl) diazoethane, to a plasmid with the desired gene–this time, the gene for luciferase, an enzyme involved in bioluminescence. When the researchers shone UV light on the vesicles, the cage quickly detached, allowing the luciferase gene to be transcribed. The researchers collected, washed, and broke open the particles with a standard cell-lysis buffer. They then added luciferin to the mixture, which reacts with luciferase to produce light. Using a luminometer, they verified that the protein was indeed functional and making light.

The researchers next did an experiment in mice, demonstrating that they could turn on protein expression in a targeted region of a living animal merely by shining light there and nowhere else. The team injected a fresh batch of luciferase-making vesicles just under the skin of the abdomens of mice and luciferin nearby. They irradiated the part of the abdomen where they’d placed the particles. An hour later, that spot glowed four times as brightly in irradiated mice as in those injected with vesicles but kept UV-free.

The study’s method for making proteins “could be extremely valuable” for drug delivery and perhaps other applications, says David Putnam of Cornell University. He is “curious to see how they plan to turn it off,” he says. If they could turn it on and off at will, doctors would be able to fine-tune drug doses. Anderson hasn’t yet come up with an off switch but would like to. He next plans to see if he can target his particles to specific cell types.