Life Built to Order

There are two main approaches to creating artificial cells, dubbed â€top down†and â€bottom up.†Genome guru Craig Venter is the most prominent top-downer. Starting with the simplest known bacterium on the planet, a harmless 517-gene organism called Mycoplasma genitalium that inhabits the human genital tract, Venter´s team at the Institute for Biological Energy Alternatives in Rockville, Maryland, is attempting to replace the organism´s natural genetic code with a stripped-down synthetic one.


That, of course, requires knowing which genes are essential to keeping the bug alive and which aren´t. By pruning genes from the bacterium one at a time, the team has already determined that as many as 215 genes might be extraneous. The next step, which the group is sweating over now, is constructing an artificial, Cliffs Notes version of the original genetic code, installing it in an organism stripped of its DNA and then seeing if they can coax this new creature to life.


Even Venter, a scientist accustomed to mega-challenges, has admitted that getting his synthetic bug to â€boot up†will be no small feat. So far, the record for a scratch-built genome is 7,500 chemical bases, assembled in 2002 by researchers at the State University of New York to create a polio virus. Venter´s bug will require a synthetic DNA strand 40 times as long. Even if his team succeeds in constructing it, nobody knows whether it´s possible to swap the entire genetic code of a living creature for an artificial version without killing the organism. Still, Venter and his colleagues are making headway. In 2003 they reported that they had sewn together a harmless virus known as phi-X (although some scientists derided the feat as little more than a publicity stunt). In the months since, Venter has been tight-lipped about his group´s progress but not about the potential promise of the Energy
Department-funded project. One possibility: loading his synthetic organism with genetic instructions to convert atmospheric carbon dioxide to methane for use as fuel.


As ambitious as Venter´s plans are, they hardly compare with what Rasmussen and other bottom-uppers are trying to accomplish, building an organism from scratch. Sitting in the living room of his adobe-style house, Rasmussen and I discuss his protocell. An impressive variety of creatures roam the grounds-horses, chickens, mosquito fish, a dog, a cat, two kids, and who knows what else. As we talk, Rasmussen´s 13-year-old son Leif appears, plucks a parakeet from a nearby cage, and places it on his dad´s shoulder. But Rasmussen is too wrapped up in the notion of creating life to pay heed to the life-form twittering into his ear.


When he began this project, he compiled a list of the minimum necessary parts for an artificial organism, then pruned it to three: a metabolism to generate energy, a DNA-like molecule to store operating instructions, and a membrane to serve as sausage casing and hold all the parts together. But he soon realized that he needed to simplify further. Even primitive single-celled organisms are works of sophisticated engineering, their membranes studded with channels for transporting nutrients in and waste out. These natural structures would be tough to duplicate.


Along with chemist Chen, who works at Argonne National Laboratory, Rasmussen pared down his design,
creating computer simulations to test his ideas as he went. â€We turned things completely upside down,†he says. Or, to be precise, inside out. For starters, Rasmussen and Chen put some of the molecular machinery on the outside of their
synthetic cell, thus doing away with the need for a fancy
channel-studded membrane. Instead the protocell is glued together by a clump of fatty-acid molecules (â€kind of like
a used wad of chewing gum,†Rasmussen explains). This molecular blob-known in the chemistry trade as a micelle -is about as primitive a membrane as you can make.


The beauty of the plan is that the micelle should assemble itself. When I ask how, Rasmussen bolts up. â€Let me show you something,†he says, scurrying off barefoot into the kitchen, where he bangs around before returning with a sloshing glass of water, a stainless-steel dish-soap dispenser, and an unreadable glint in his ice-blue eyes. I realize that he´s about to perform an impromptu experiment-and I briefly wonder if I should duck for cover. Rasmussen has confessed that he´s more comfortable at a computer than a chemistry bench. Years ago Peter Nielsen, the Danish chemist who is collaborating on the protocell project, invited Rasmussen to his lab to â€get his hands wet,†as experimentalists like to
say. Rasmussen did, and then some. Neither man will reveal precisely what happened, just that it involved the accidental spilling of a certain radioactive substance and the ruining of an experiment. In fact, when Rasmussen first asked Nielsen to help with the protocell, Nielsen agreed, but only after joking that Rasmussen must promise not to touch anything.


Back in Rasmussen´s living room, I watch him pump a few shots of soap into the water glass, cup his hand over the rim, and shake as if he were mixing a cocktail. Liquid erupts from between his fingers, splattering his shirt. Rasmussen curses under his breath, then holds out the glass for me to inspect. Delicate bubbles swirl in the cloudy water. OK, so?


So, he explains, looking slightly deflated that his Mr. Wizard bit didn´t suffice, soap is what chemists call a surfactant. On a sheet of paper, he sketches something that looks like
a sperm. The head of the soap molecule, he explains, is attracted to water, the tail repelled by it. Spritz enough of these part-hydrophilic, part-hydrophobic structures into water, and the molecules automatically ball up into micelles. Rasmussen and his team won´t use dish soap, of course, but some other surfactant; like many details of the protocell´s design, they will know which one only after intensive experimentation. But the basic recipe for protocells starts with throwing a fatty-acid surfactant into a beaker of water. In the blink of an eye, there should be, as Rasmussen puts it,
â€zillions†of blobby micelles swirling inside.


Next, the genetic material. Most organisms operate with DNA or RNA. But Rasmussen and his group plan to try a man-made nucleic acid called PNA, or peptide nucleic acid. Synthesized by Nielsen and his colleagues in the early 1990s, PNA looks and acts much like DNA-same double-helix shape, same four chemical bases. But rather than a backbone composed of sugar-phosphate molecules, PNA has one made of peptides, the building blocks of proteins.


Rasmussen´s PNA-based protocell might help solve a long-standing riddle: What was the ur-gene? One leading theory is that the earliest organism relied on a self-replicating version of RNA. But in 2000, Stanley Miller, the father of
origins-of-life research, suggested that PNA ingredients were also present on the early Earth. Could the first life-form
have been weirder than we thought, a PNA-based creature? Rasmussen´s protocell will test that notion.


The main advantage of PNA, though, is that it is electrically conductive, so in addition to acting as genetic material, it jump-starts the protocell´s metabolism. In the initial blueprint, a photosensitive molecule-the alcohol pinacol is one option-and short PNA strands are thrown into the mix [see illustration]. After running a series of simulations in November, Rasmussen and Chen conceived of attaching the pinacol to the ends of PNA strands before throwing it into the beaker. When light strikes the pinacol, it will cause the compound to throw off an electron, which will streak down the PNA bases. When it reaches the other end, scientists expect it to trigger a chemical reaction with the final ingredient they plan to throw into the mix: food.


The food consists of precursor molecules that the protocell´s PNA-pinacol metabolism will convert into new fatty acids and PNA molecules. Without the addition of these precursors, Rasmussen explains, the protocells â€would pretty much just sit there doing nothing.†The newly created fatty acids will be incorporated into existing micelles, causing them to grow until they become unstable and pinch in two-protocell procreation. An adult protocell will measure merely five to 10 nanometers across; in comparison, M. genitalium, the organism Venter and his team are working with, is between 200 and 250 nanometers. â€We couldn´t imagine anything that´s simpler,†Rasmussen says.


With only three basic parts, the protocell itself may be simple, but the chemistry that brings it to life is wildly complex. On paper, at least, the micelles should soak up the precursor molecules, providing a ready store of â€food†for the light-powered metabolism to act on.
Single-stranded PNA molecules, meanwhile, should cling to the micelle´s
exterior and pair with complementary strands of PNA that have been created by the organism´s own metabolism. But who knows? Rasmussen says it remains to be seen how all these molecules will actually behave in solution. â€If we really knew ahead of time how to do this,†notes William â€Woody†Woodruff, a Los Alamos chemist on Rasmussen´s team, â€we would have created life already.â€

Some of the experts who have seen Rasmussen´s blueprint have serious doubts that this Rube Goldbergian organism will work. When Rasmussen talks about his protocells at astrobiology conferences and other such gatherings, his work isn´t always warmly received. The words â€abstract†and â€weird†pop up a lot. â€It´s too far-fetched,†argues chemist Pier Luigi Luisi of the University of Rome. Luisi says he wants to see experimental data before he buys into Rasmussen´s approach: â€You cannot convince anybody with calculations on a blackboard.â€

Other scientists emphasize that it´s not yet clear what route-top-down, bottom-up or something in between-may ultimately lead to artificial life, so it would be premature to dismiss Rasmussen. Biophysicist Andrew Pohorille of the NASA Ames Research Center in California contends that Rasmussen has as much chance of creating life as anybody-maybe more, since few researchers out there have put as much thought into it: â€He is definitely way ahead of the curve.â€Rasmussen thinks his critics exaggerate. â€It sounds mind-boggling,†he says, â€mostly because we have a pretty rigid idea of what life is.†He won the three-year Los Alamos grant in part because of computer models he devised showing that the protocell could work. More important, he and Chen have been able to show in the lab that their primitive light-sensitive meta-bolism can create the chewing-gum-like membrane molecules.


Meanwhile, Rasmussen is exploring other opportunities. He and several scientists from the European artificial-cell effort have formed a Venice, Italy-based startup called ProtoLife. Last June, Rasmussen flew to Silicon Valley with two partners to speak with potential investors. One commercial possibility
Rasmussen envisions is turning protocells into drug-delivery
vehicles. The protocell, he says, could be designed to sense when it encounters a particular type of tissue in the body and then to dump its cargo. Rasmussen also imagines making protocells that could withstand high toxicity levels. Such â€Terminator†cells could be used to sop up nasty contaminants such as perchlorate, something existing remediation systems don´t do well.


If you really get Rasmussen going, he´ll rhapsodize about far-out stuff like self-healing coatings for aircraft. And why not? All organisms have evolved mechanisms to repair themselves, and Rasmussen thinks that protocells should ultimately be capable of doing the same, opening the door to all kinds of exotic applications. True, a more practical approach to such problems would be to fiddle with the DNA of existing organisms-as many scientists are already doing. But Rasmussen says that the option to build entirely new creatures will give scientists a
fresh palette. Nevertheless, it seems premature to be articulating applications. The venture capitalists whom Rasmussen
approached apparently agreed; they declined to cut a check.
One day when Rasmussen was busy, I asked someone in the Los Alamos public relations office to show me the lab where the first artificial life-form might be created. Driving on a private road, my guide, Nancy, and I passed TA-55, the place where
plutonium is sculpted into grapefruit-size bulbs for thermonuclear bombs. Nancy remarked that it´s the most heavily guarded complex on the mesa. Then she looked in her rearview mirror and saw that a white SUV with government plates was driving close behind. Not until we pulled into the driveway of the lab where the protocell work would take place did it peel away.


Even when you´re not being followed, it´s impossible to
wander Los Alamos, where street signs carry names like Bikini Atoll Road and Trinity Drive, without pondering the double-sided nature of novel technologies or the irony that a place identified with the most fearsome killing machine in history might one day spawn a new kind of life. For now, though, Rasmussen says he´s just concerned with getting something to happen inside his team´s shiny glass beakers.


â€Just one fricking life cycle,†he tells me later, sliding his hand through his hair, â€would be wonderful.â€