Biologist Andrei Sakai has grown ‘liters and liters of those cells’ and then destroyed them

How did life on Earth originate? How did the first cells form, what did they look like, and how did they work? Chemical biologist Andrei Sakai searched for answers for almost five years – including a six-month ‘corona delay’. In vain, it seemed. Because one experiment after another failed. Until, at the end of his PhD research, he still made important breakthroughs.

On December 4, he received his PhD from Radboud University under Professor Wilhelm Huck, who won the Spinoza Prize in 2016 for his research into the origins of life. Sakai: “We gave up my project twice. But then we got new ideas and we picked it up again. You have to persevere.”

Huck’s group approaches the problem from two sides: they remove parts from the most basic unit of life, the cell, and see what happens – the top-down method. And from the other side, bottom-up, they build individual molecules into more complex units. Until finally something is created that is alive.

“I combined the two,” says Sakai in a conference room, behind an open laptop. “I started with simple bacterial cells, broke them down, separated the main parts, and then put them back together in the hope that cells would form again.” If he succeeded, he would be the first scientist to create a synthetic cell bottom-up. The idea that such a cell can be made in the lab would then be proven.

For his research, Sakai used so-called JCVI-syn3A cells, processed cells from Mycoplasma-bacteria. He received this from the J. Craig Venter Institute in Rockville, with which Huck had started a collaboration at the end of 2018. The founder of that institute, biotechnologist and businessman John Craig Venter, had made a name for himself in 2001 by also the firstto publish the rough code of a human genome (Science, February 16, 2001). Fifteen years later he caused a stir again by creating the first (top-down) synthetic cell, based on Mycoplasma-bacteria. Mycoplasma is one of the simplest life forms that can grow on its own (unlike viruses, which require a host). They simplified that synthetic cell (JCVI-syn1.0) at the Venter Institute by removing genes from it. This ultimately led to the JCVI-syn3A cell, which Sakai used for his experiments.

A soup with proteins

“I have grown liters and liters of those cells. I then exploded the cells. You can then separate the three main parts: membrane, DNA and cytoplasm. The cytoplasm is a soup of proteins.”

Sakai then examined those parts separately. “Because we speculated that no cell would be created if we put the parts together like this.” For example, his research showed that the DNA was very damaged. A slide explaining the process appears on his laptop. “This one slide is half of my PhD,” he smiles. “When purifying the DNA, you have to go through a number of steps, in which you put the DNA with some liquids in a small tube, a tube. You mix it by sucking everything in with a pipette a few times and spitting it out again. You do a lot of damage with all that pipetting. The DNA breaks. We managed to prevent this by combining different methods. You add agarose. This forms a protective covering around the DNA, so that a significant part remains intact.”

Something also appeared to be wrong with the cytoplasm. Sakai discovered increased activity of an enzyme that breaks down RNA – a cell translates pieces of DNA into RNA codes, on the basis of which it builds proteins. “Mycoplasmas are parasites. On their outside they produce a lot of these enzymes, ribonucleases,” Sakai explains. In a host, they break down the RNA into building blocks, nucleosides, which the mycoplasmas take up to build their own hereditary material. “The worst part was that in our isolated cytoplasm even the ribosomes were broken down.” These complexes of proteins and RNA play a central role in the production of proteins.

Cells exploding

“I have tried everything to solve this,” says Sakai. “For two and a half years we only had negative results. I ended up trying a method that we thought wouldn’t work. But you try anyway. With this method we place cells in a pressure vessel with nitrogen gas. You first increase the pressure, causing the gas to enter the cells. If you then quickly release the pressure, the gas suddenly expands and the cells explode. But now it turned out that there were far fewer ribonucleases in the cytoplasm. It worked. That was like…” Sakai throws his hands in the air. “…Haaah. The breakthrough!”

This seems to open the way to the first bottom-up synthetic cell again. “I will start those experiments soon,” says Sakai, who is now a postdoc in Huck’s group.

He has been interested in the basics of life for a long time, he says. “I like working on the border between chemistry and biology. It also teaches you a lot about yourself, about your cells, how you are put together. The complexity of biological systems fascinates me. As a child I would completely take apart flowers from our garden.”

“And I also really like building things from scratch. I used to build a lot with Lego. I still do, by the way, together with my wife, who works as an engineer at Leiden University. Our most beautiful and largest Lego building is Hogwarts, the castle from the Harry Potter stories. It took us a few days to do that.”

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