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  • richardmitnick 9:17 am on November 1, 2018 Permalink | Reply
    Tags: , , , , The Baker Lab U Washington,   

    From Discover Magazine: “Meet the Biochemist Engineering Proteins From Scratch” 

    DiscoverMag

    From Discover Magazine

    October 30, 2018
    Jonathon Keats

    1
    David Baker. Brian Dalbalcon/UW Medicine

    U Washington Dr. David Baker

    In a sleek biochemistry laboratory at the University of Washington, postdoctoral fellow Yang Hsia is watching yellowish goo — the liquefied remains of E. coli — ooze through what looks like a gob of white marshmallow. “This isn’t super exciting,” he says.

    While growing proteins in bacteria and then purifying them, using blobby white resin as a filter, doesn’t make for riveting viewing, the end product is extraordinary. Accumulating in Hsia’s resin is a totally artificial protein, unlike anything seen in nature, that might just be the ideal chassis for the first universal flu vaccine.

    David Baker, Hsia’s adviser, calls this designer protein a “Death Star.” Imaged on his computer, its structure shows some resemblance to the notorious Star Wars superweapon. Though microscopic, by protein standards it’s enormous: a sphere made out of many interlocking pieces.

    2
    The Death Star artificial protein. Institute for Protein Design

    “We’ve figured out a way to put these building blocks together at the right angles to form these very complex nanostructures,” Baker explains. He plans to stud the exterior with proteins from a whole suite of flu strains so that the immune system will learn to recognize them and be prepared to fend off future invaders. A single Death Star will carry 20 different strains of the influenza virus.

    Baker hopes this collection will cover the entire range of possible influenza mutation combinations. This all-in-one preview of present and future flu strains could replace annual shots: Get the Death Star vaccination, and you’ll already have the requisite antibodies in your bloodstream.

    As Baker bets on designer proteins to defeat influenza, others are betting on David Baker.

    After revolutionizing the study of proteins — molecules that perform crucial tasks in every cell of every natural organism — Baker is now engineering them from scratch to improve on nature. In late 2017, the Open Philanthropy Project gave his University of Washington Institute for Protein Design more than $10 million to develop the Death Star and support Rosetta, the software platform he conceived in the 1990s to discover how proteins are assembled. Rosetta has allowed Baker’s lab not only to advance basic science and pioneer new kinds of vaccines, but also to create drugs for genetic disorders, biosensors to detect toxins and enzymes to convert waste into biofuels.

    His team currently numbers about 80 grad students and postdocs, and Baker is in constant contact with all of them. He challenges their assumptions and tweaks their experiments while maintaining an egalitarian environment in which ideas may come from anyone. He calls his operation a “communal brain.” Over the past quarter-century, this brain has generated nearly 450 scientific papers.

    “David is literally creating a new field of chemistry right in front of our eyes,” says Raymond Deshaies, senior vice president for discovery research at the biotech company Amgen and former professor of biology at Caltech. “He’s had one first after another.”

    Nature’s Origami

    When Baker was studying philosophy at Harvard University, he took a biology class that taught him about the so-called “protein folding problem.” The year was 1983, and scientists were still trying to make sense of an experiment, carried out in the early ’60s by biochemist Christian Anfinsen, that revealed the fundamental building blocks of all life on Earth were more complex than anyone imagined.

    The experiment was relatively straightforward. Anfinsen mixed a sample of the protein ribonuclease — which breaks down RNA — with a denaturant, a chemical that deactivated it. Then he allowed the denaturant to evaporate. The protein started to function again as if nothing ever happened.

    What made this simple experiment so striking was the fact that the amino acids in protein molecules are folded in three-dimensional forms that make origami look like child’s play. When the denaturant unfolded Anfinsen’s ribonuclease, there were myriad ways it could refold, resulting in structures as different as an origami crane and a paper airplane. Much as the folds determine whether a piece of paper can fly across a room, only one fold pattern would result in functioning ribonuclease. So the puzzle was this: How do proteins “know” how to refold properly?

    “Anfinsen showed that the information for both structure and activity resided in the sequence of amino acids,” says University of California, Los Angeles, biochemist David Eisenberg, who has been researching protein folding since the 1960s. “There was a hope that it would be possible to use sequence information to get three-dimensional structural information. Well, that proved much more difficult than anticipated.”

    2
    Protein molecules play critical roles in every aspect of life. The way each protein folds determines its function, and the ways to fold are virtually limitless, as shown in this small selection of proteins visualized through the software platform Rosetta, born in Baker’s lab. Institute for Protein Design.

    Baker was interested enough in protein folding and other unsolved mysteries of biology to switch majors and apply to grad school. “I’d never worked in a lab before,” he recalls. He had only a vague notion of what biologists did on a daily basis, but he also sensed that the big questions in science, unlike philosophy, could actually be answered.

    Grad school plunged Baker into the tediousness and frustrations of benchwork, while also nurturing some of the qualities that would later distinguish him. He pursued his Ph.D. under Randy Schekman, who was studying how molecules move within cells, at the University of California, Berkeley. To aid in this research, students were assigned the task of dismantling living cells to observe their internal molecular traffic. Nearly half a dozen of them, frustrated by the assignment’s difficulty, had given up by the time Baker got the job.

    Baker decided to follow his instincts even though it meant going against Schekman’s instructions. Instead of attempting to keep the processes within a cell still functioning as he dissected it under his microscope, Baker concentrated on preserving cell structure. If the cell were a wristwatch, his approach would be equivalent to focusing on the relationship between gears, rather than trying to keep it ticking, while taking it apart.

    “He was completely obsessed,” recalls Deshaies, who was his labmate at the time (and one of the students who’d surrendered). Nobody could stop Baker, or dissuade him. He worked for months until he proved his approach was correct: Cell structure drove function, so maintaining its anatomy preserved the internal transportation network. Deshaies believes Baker’s methodological breakthrough was “at the core of Randy’s Nobel Prize,” awarded in 2013 for working out one of the fundamentals of cellular machinery.

    But Baker didn’t dwell on his achievement, or cell biology for that matter. By 1989, Ph.D. in hand, he’d headed across the Bay to the University of California, San Francisco, where he switched his focus to structural biology and biochemistry. There he built computer models to study the physical properties of the proteins he worked with at the bench. Anfinsen’s puzzle remained unsolved, and when Baker got his first faculty appointment at the University of Washington, he took up the protein-folding problem full time.

    From Baker’s perspective, this progression was perfectly natural: “I was getting to more and more fundamental problems.” Deshaies believes Baker’s tortuous path, from cells to atoms and from test tubes to computers, has been a factor in his success. “He just has greater breadth than most people. And you couldn’t do what he’s done without being somewhat of a polymath.”

    3
    Illustration above: National Science foundation. Illustrations below: Jay Smith

    Rosetta Milestone

    Every summer for more than a decade, scores of protein-folding experts convene at a resort in Washington’s Cascade Mountains for four days of hiking and shop talk. The only subject on the agenda: how to advance the software platform known as Rosetta.

    David Baker’s Rosetta@home project, a project running on BOINC software from UC Berkeley


    Rosetta@home BOINC project



    They call it Rosettacon.

    Rosetta has been the single most important tool in the quest to understand how proteins fold, and to design new proteins based on that knowledge. It is the link between Anfinsen’s ribonuclease experiment and Baker’s Death Star vaccine.

    When Baker arrived at the University of Washington in 1993, researchers knew that a protein’s function was determined by its structure, which was determined by the sequence of its amino acids. Just 20 different amino acids were known to provide all the raw ingredients. (Their particular order — specified by DNA — makes one protein fold into, say, a muscle fiber and another fold into a hormone.) Advances in X-ray crystallography, a technique for imaging molecular structure, had provided images of many proteins in all their folded splendor. Sequencing techniques had also improved, benefitting from the Human Genome Project as well as the exponential increase in raw computing power.

    “There’s a right time for things,” Baker says in retrospect. “To some extent, it’s just luck and historical circumstance. This was definitely the right time for this field.”

    Which is not to say that modeling proteins on a computer was a simple matter of plugging in the data. Proteins fold to their lowest free energy state: All of their amino acids must align in equilibrium. The trouble is that the equilibrium state is just one of hundreds of thousands of options — or millions, if the amino acid sequence is long. That’s far too many possibilities to test one at a time. Nature must have another way of operating, given that folding is almost instantaneous.

    Baker’s initial approach was to study what nature was doing. He broke apart proteins to see how individual pieces behaved, and he found that each fragment was fluctuating among many possible structures. “And then folding would occur when they all happened to be in the right geometry at the same time,” he says. Baker designed Rosetta to simulate this dance for any amino acid sequence.

    Baker wasn’t alone in trying to predict how proteins fold. In 1994, the protein research community organized a biennial competition called CASP (Critical Assessment of Protein Structure Prediction). Competitors were given the amino acid sequences of proteins and challenged to anticipate how they would fold.

    The first two contests were a flop. Structures that competitors number-crunched looked nothing like folded proteins, let alone the specific proteins they were meant to predict. Then everything changed in 1998.

    3
    Rosetta’s impressive computational power allows researchers to predict how proteins — long, complex chains of amino acids — will fold; the platform also helps them reverse engineer synthetic proteins to perform specific tasks in medicine and other fields. Brian Dalbalcon/UW Medicine.

    Function Follows Form

    That summer, Baker’s team received 20 sequences from CASP, a considerable number of proteins to model. But Baker was optimistic: Rosetta would transform protein-folding prediction from a parlor game into legitimate science.

    In addition to incorporating fresh insights from the bench, team members — using a janky collection of computers made of spare parts — found a way to run rough simulations tens of thousands of times to determine which fold combinations were most likely.

    They successfully predicted structures for 12 out of the 20 proteins. The predictions were the best yet, but still approximations of actual proteins. In essence, the picture was correct, but blurry.

    Improvements followed rapidly, with increased computing power contributing to higher-resolution models, as well as improved ability to predict the folding of longer amino acid chains. One major leap was the 2005 launch of Rosetta@Home, a screensaver that runs Rosetta on hundreds of thousands of networked personal computers whenever they’re not being used by their owners.

    Yet the most significant source of progress has been RosettaCommons, the community that has formed around Rosetta. Originating in Baker’s laboratory and growing with the ever-increasing number of University of Washington graduates — as well as their students and colleagues — it is Baker’s communal brain writ large.

    Dozens of labs continue to refine the software, adding insights from genetics and methods from machine learning. New ideas and applications are constantly emerging.

    4
    Protein (in green) enveloping fentanyl molecule. Bick et al. eLife 2017.

    The communal brain has answered Anfinsen’s big question — a protein’s specific amino acid alignment creates its unique folding structure — and is now posing even bigger ones.

    “I think the protein-folding problem is effectively solved,” Baker says. “We can’t necessarily predict every protein structure accurately, but we understand the principles.

    “There are so many things that proteins do in nature: light harvesting, energy storage, motion, computation,” he adds. “Proteins that just evolved by pure, blind chance can do all these amazing things. What happens if you actually design proteins intelligently?”

    De Novo Design

    Matthew Bick is trying to coax a protein into giving up its sugar habit for a full-blown fentanyl addiction. His computer screen shows a colorful image of ribbons and swirls representing the protein’s molecular structure. A sort of Technicolor Tinkertoy floats near the center, representing the opioid. “You see how it has really good packing?” he asks me, tracing the ribbons with his finger. “The protein kind of envelops the whole fentanyl molecule like a hot dog bun.”

    A postdoctoral fellow in Baker’s lab, Bick engineers protein biosensors using Rosetta. The project originated with the U.S. Department of Defense. “Back in 2002, Chechen rebels took a bunch of people hostage, and there was a standoff with the Russian government,” he says. The Russians released a gas, widely believed to contain a fentanyl derivative, that killed more than a hundred people. Since then, the Defense Department has been interested in simple ways to detect fentanyl in the environment in case it’s used for chemical warfare in the future.

    Proteins are ideal molecular sensors. In the natural world, they’ve evolved to bind to specific molecules like a lock and key. The body uses this system to identify substances in its environment. Scent is one example; specific volatiles from nutrients and toxins fit into dedicated proteins lining the nose, the first step in alerting the brain to their presence. With protein design, the lock can be engineered to order.

    For the fentanyl project, Bick instructed Rosetta to modify a protein with a natural affinity for the sugar xylotetraose. The software generated hundreds of thousands of designs, each representing a modification of the amino acid sequence predicted to envelop fentanyl instead of sugar molecules. An algorithm then selected the best several hundred options, which Bick evaluated by eye, eventually choosing 62 promising candidates. The protein on Bick’s screen was one of his favorites.

    “After this, we do the arduous work of testing designs in the lab,” Bick says.

    5
    Cassie Bryan, a senior fellow at Baker’s Institute for Protein Design at the University of Washington, checks on a tube of synthetic proteins. The proteins, not seen in nature, are in the process of thawing and being prepped to test how they perform. Brian Dalbalcon/UW Medicine.

    With another image, he reveals his results. All 62 contenders have been grown in yeast cells infused with synthetic genes that spur the yeasts’ own amino acids to produce the foreign proteins. The transgenic yeast cells have been exposed to fentanyl molecules tagged with a fluorescing chemical. By measuring the fluorescence — essentially shining ultraviolet light on the yeast cells to see how many glow with fentanyl — Bick can determine which candidates bind to the opioid with the greatest strength and consistency.

    Baker’s lab has already leveraged this research to make a practical environmental sensor. Modified to glow when fentanyl binds to the receptor site, Bick’s customized protein can now be grown in a common plant called thale cress. This transgenic weed can cover terrain where chemical weapons might get deployed, and then glow if the dangerous substances are present, providing an early warning system for soldiers and health workers.

    The concept can also be applied to other biohazards. For instance, Bick is now developing a sensor for aflatoxin, a residue of fungus that grows on grain, causing liver cancer when consumed by humans. He wants the sensor to be expressed in the grain itself, letting people know when their food is unsafe.

    But he’s going about things differently this time around. Instead of modifying an existing protein, he’s starting from scratch. “That way, we can control a lot of things better than in natural proteins,” he explains. His de novo protein can be much simpler, and have more predictable behavior, because it doesn’t carry many million years of evolutionary baggage.

    For Baker, de novo design represents the summit of his quarter-century quest. The latest advances in Rosetta allow him to work backward from a desired function to an appropriate structure to a suitable amino acid sequence. And he can use any amino acids at all — thousands of options, some already synthesized and others waiting to be designed — not only the 20 that are standard in nature for building proteins.

    Without the freedom of de novo protein design, Baker’s Death Star would never have gotten off the ground. His group is now also designing artificial viruses. Like natural viruses, these protein shells can inject genetic material into cells. But instead of infecting you with a pathogen, their imported DNA would patch dangerous inherited mutations. Other projects aim to take on diseases ranging from malaria to Alzheimer’s.

    In Baker’s presence, protein design no longer seems so extraordinary. Coming out of a brainstorming session — his third or fourth of the day — he pulls me aside and makes the case that his calling is essentially the destiny of our species.

    “All the proteins in the world today are the product of natural selection,” he tells me. “But the current world is quite a bit different than the world in which we evolved. We live much longer, so we have a whole new class of diseases. We put all these nasty chemicals into the environment. We have new needs for capturing energy.

    “Novel proteins could solve a lot of the problems that we face today,” he says, already moving to his next meeting. “The goal of protein design is to bring those into existence.”

    See the full article here .

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  • richardmitnick 6:52 pm on March 6, 2018 Permalink | Reply
    Tags: , , , , , The Baker Lab U Washington, U Wasington Medicine News Room   

    From UW Medicine Newsroom: “Scientists create complex transmembrane proteins from scratch” 

    U Washington
    University of Washington

    UW Medicine Newsroom

    March 1, 2018
    Leila Gray
    206.685.0381
    leilag@uw.edu

    The ability to build transmembrane proteins opens the way for custom-designing structures that span living cell membranes and perform new tasks.

    1
    Four computer-designed proteins combine to form a transmembrane tetramer with the top of structure facing the cytoplasm. Institute for Protein Design.

    In the living world, transmembrane proteins are found embedded in the membrane of all cells and cellular organelles. They are essential for them to function normally. For example, many naturally occurring transmembrane proteins act as gateways for the movement of specific substances across a biological membrane. Some transmembrane proteins receive or transmit cell signals. Because of such roles, many drugs are designed to target transmembrane proteins and alter their function.

    Now researcher are looking at designing the transmembrane proteins themselve to perform specific tasks.

    “Our results pave the way for the design of multispan membrane proteins that could mimic proteins found in nature or have entirely novel structure, function and uses,” said David Baker, a University of Washington School of Medicine professor biochemistry and director of the UW Institute of Protein Design who led the project.

    U Washington Dr. David Baker

    David Baker’s Rosetta@home project, a project running on BOINC software from UC Berkeley

    But understanding how transmembrane proteins are put together and how they work has proved challenging. Because they act while embedded within the cellular membrane, transmembrane proteins have proven to be more difficult to study than proteins that operate in the watery solution that make up the cells’ cytoplasm or in the extracellular fluid.

    In the new study, Lu and his coworkers used a computer program, developed in the Baker lab and called Rosetta, that can predict the structure a protein will fold into after it has been synthesized. The architecture of a protein is crucial because a protein’s structure determines its function.

    A protein’s shape forms from complex interactions between the amino acids that make up the protein chain and between the amino acids and the surrounding environment. Ultimately, the protein assumes the shape that best balances out all these factors so that the protein achieves the lowest possible energy state.

    The Rosetta program used by Lu and his colleagues can predict the structure of a protein by taking into account these interactions and calculating the lowest overall energy state. It is not unusual for the program to create tens of thousands of model structures for an amino acid sequence and then identify the ones with lowest energy state. The resulting models have been shown to accurately represent the structure the sequence will likely assume in nature.

    Determining the structure of transmembrane proteins is difficult because portions of transmembrane proteins must pass though the membrane’s interior, which is made of oily fats called lipids.

    In aqueous fluids, amino acid residues that have polar sidechains – components that can have a charge under certain physiological conditions or that participate in hydrogen bonding — tend to be located on the surface of the protein where they can interact with water, which has negatively and positively side charges to its molecule. As a result, polar residues on proteins are called hydrophilic, or “water-loving.”

    Non-polar residues, on the other hand, tend to be found packed within the protein core away from the polar aqueous fluid. Such residues are called hydrophobic or “water-fearing.” As a result, the interaction between the water-loving and water-fearing residues of the protein and the surrounding watery fluids helps drive protein folding and stabilizes the protein’s final structure.

    In membranes, however, protein folding is more complicated because the lipid interior of the membrane is non-polar, that is, it has no separation of electrical charges. This means to be stable the protein must place nonpolar, water-fearing residues on its surface, and pack its polar, water-loving residues inside. Then it must find a way to stabilize its structure by creating bonds between the hydrophilic residues within its core.

    The key to solving the problem, said Lu, was to apply a method developed by Baker lab to design the transmembrane portion so that the polar, hydrophilic residues fit in such a way that enough would form hydrongen bonds– that can tie the protein together from within

    “Putting together these ‘buried hydrogen bond networks’ was like putting together a jig-saw puzzle,” Baker said.

    With this approach, Lu and his colleagues were able to manufacture the designed transmembrane proteins inside bacteria and mammalian cells by using as many as 215 amino acids. The resulting proteins proved to be highly thermally stable and able to correctly orient themselves on the membrane. Like naturally occurring transmembrane proteins, the proteins are multipass, meaning they traverse the membrane several times, and assemble into stable multi-protein complexes, such as dimers, trimers and tetramers.

    “We have shown that it is now possible to accurately design complex, multipass transmembrane proteins that can be expressed in cells. This will make it possible for researchers to design transmembrane proteins with entirely novel structures and functions,” said Lu.

    This work was supported by the Howard Hughes Medical Institute, National Institutes of Health (R01GM063919), the Raymond and Beverly Sackler fellowship, and the National Research Foundation of Korea (NRF- 2016R1A6A3A03007871).

    The research is reported in the March 1 issue of the journal Science. Peilong Lu, a senior fellow in the Baker lab, is the paper’s lead author.

    See the full article here .

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  • richardmitnick 5:20 pm on January 2, 2018 Permalink | Reply
    Tags: , , , , , Scientists Are Designing Artisanal Proteins for Your Body, The Baker Lab U Washington   

    From NYT: “Scientists Are Designing Artisanal Proteins for Your Body” 

    New York Times

    The New York Times

    DEC. 26, 2017
    CARL ZIMMER

    1
    John Hersey

    The human body makes tens of thousands of cellular proteins, each for a particular
    task. Now researchers have learned to create custom versions not found in nature.

    Our bodies make roughly 20,000 different kinds of proteins, from the collagen in our skin to the hemoglobin in our blood. Some take the shape of molecular sheets. Others are sculpted into fibers, boxes, tunnels, even scissors.

    A protein’s particular shape enables it to do a particular job, whether ferrying oxygen through the body or helping to digest food.

    Scientists have studied proteins for nearly two centuries, and over that time they’ve worked out how cells create them from simple building blocks. They have long dreamed of assembling those elements into new proteins not found in nature.

    But they’ve been stumped by one great mystery: how the building blocks in a protein take their final shape. David Baker, 55, the director of the Institute for Protein Design at the University of Washington, has been investigating that enigma for a quarter-century.

    Now, it looks as if he and his colleagues have cracked it. Thanks in part to crowdsourced computers and smartphones belonging to over a million volunteers, the scientists have figured out how to choose the building blocks required to create a protein that will take on the shape they want.

    In a series of papers published this year, Dr. Baker and his colleagues unveiled the results of this work. They have produced thousands of different kinds of proteins, which assume the shape the scientists had predicted. Often those proteins are profoundly different from any found in nature.

    This expertise has led to a profound scientific advance: cellular proteins designed by man, not by nature. “We can now build proteins from scratch from first principles to do what we want,” said Dr. Baker.

    2
    Dr. David Baker in his lab at the University of Washington, where scientists are learning how to create cellular proteins to perform a variety of tasks. Credit Evan McGlinn for The New York Times.

    Scientists soon will be able to construct precise molecular tools for a vast range of tasks, he predicts. Already, his team has built proteins for purposes ranging from fighting flu viruses to breaking down gluten in food to detecting trace amounts of opioid drugs.

    William DeGrado, a molecular biologist at the University of California, San Francisco, said the recent studies by Dr. Baker and his colleagues represent a milestone in this line of scientific inquiry. “In the 1980s, we dreamed about having such impressive outcomes,” he said.

    Every protein in nature is encoded by a gene. With that stretch of DNA as its guide, a cell assembles a corresponding protein from building blocks known as amino acids.

    Selecting from twenty or so different types, the cell builds a chain of amino acids. That chain may stretch dozens, hundreds or even thousands of units long. Once the cell finishes, the chain folds on itself, typically in just a few hundredths of a second.

    Proteins fold because each amino acid has an electric charge. Parts of the protein chain are attracted to one another while other parts are repelled. Some bonds between the amino acids will yield easily under these forces; rigid bonds will resist.

    The combination of all these atomic forces makes each protein a staggering molecular puzzle. When Dr. Baker attended graduate school at the University of California, Berkeley, no one knew how to look at a chain of amino acids and predict the shape into which it would fold. Protein scientists referred to the enigma simply as “the folding problem.”

    The folding problem left scientists in the Stone Age when it came to manipulating these important biological elements. They could only use proteins that they happened to find in nature, like early humans finding sharp rocks to cut meat from bones.

    We’ve used proteins for thousands of years. Early cheese makers, for example, made milk curdle by adding a piece of calf stomach to it. The protein chymosin, produced in the stomach, turned liquid milk into a semisolid form.

    Today scientists are still looking for ways to harness proteins. Some researchers are studying proteins in abalone shells in hopes of creating stronger body armor, for instance. Others are investigating spider silk for making parachute cords. Researchers also are experimenting with modest changes to natural proteins to see if tweaks let them do new things.

    To Dr. Baker and many other protein scientists, however, this sort tinkering has been deeply unsatisfying. The proteins found in nature represent only a minuscule fraction of the “protein universe” — all the proteins that could possibly be made with varying combinations of amino acids.

    “When people want a new protein, they look around in nature for things that already exist,” Dr. Baker said. “There’s no design involved.”

    Crowdsourced Discovery

    Dr. Baker has an elfin face, a cheerful demeanor, hair that can verge on chaotic, and a penchant for wearing T-shirts to scientific presentations. But his appearance belies a relentless drive.

    After graduating from Berkeley and joining the University of Washington, Dr. Baker joined the effort to solve the folding problem. He and his colleagues took advantage of the fact that natural proteins are somewhat similar to one another.

    New proteins do not just pop into existence; they all evolve from ancestral proteins. Whenever scientists figured out the shape of a particular protein, they were able to make informed guesses about the shapes of related ones.

    Scientists also relied on the fact that many proteins are made of similar parts. One common feature is a spiral stretch of amino acids called an alpha helix. Researchers learned how to recognize the series of amino acids that fold into these spirals.

    3
    John Hersey

    In the late 1990s, the team at the University of Washington turned to software for individual studies of complex proteins. The lab decided to create a common language for all this code, so that researchers could access the collective knowledge about proteins.

    In 1998, they launched a platform called Rosetta, which scientists use to build virtual chains of amino acids and then compute the most likely form they will fold into.

    A community of protein scientists, known as the Rosetta Commons, grew around the platform. For the past twenty years, they’ve been improving the software on a daily basis and using it to better understand the shape of proteins — and how those shapes enable them to work.

    In 2005, Dr. Baker launched a program called Rosetta@home, which recruited volunteers to donate processing time on their home computers and, eventually, Android phones. Over the past 12 years, 1,266,542 people have joined the Rosetta@home community.

    My BOINC

    I have 1,005,660 BOINC credits for Rosetta from my days as a BOINC cruncher.

    Rosetta@home project, a project running on BOINC software from UC Berkeley


    Step by step, Rosetta grew more powerful and more sophisticated, and the scientists were able to use the crowdsourced processing power to simulate folding proteins in greater detail. Their predictions grew startlingly more accurate.

    The researchers went beyond proteins that already exist to proteins with unnatural sequences. To see what these unnatural proteins looked like in real life, the scientists synthesized genes for them and plugged them into yeast cells, which then manufactured the lab’s creations.

    “There are subtleties going on in naturally occurring proteins that we still don’t understand,” Dr. Baker said. “But we’ve mostly solved the folding problem.”

    Proteins and Pandemics

    These advances gave Dr. Baker’s team the confidence to take on an even bigger challenge: They began to design proteins from scratch for particular jobs. The researchers would start with a task they wanted a protein to do, and then figure out the string of amino acids that would fold the right way to get the job done.

    In one of their experiments, they teamed up with Ian Wilson, a virologist at Scripps Research Institute, to devise a protein to fight the flu.

    Dr. Wilson has been searching ways to neutralize the infection, and his lab had identified one particularly promising target: a pocket on the surface of the virus. If scientists could make a protein that fit snugly in that pocket, it might prevent the virus from slipping into cells.

    Dr. Baker’s team used Rosetta to design such a protein, narrowing their search to several thousand of chains of amino acids that might do the job. They simulated the folding of each one, looking for the combinations that might fit into the viral niche.

    The researchers then used engineered yeast to turn the semifinalists into real proteins. They turned the proteins loose on the flu viruses. Some grabbed onto the viruses better than others, and the researchers refined their molecular creations until they ended up with one they named HB1.6928.2.3.

    To see how effective HB1.6928.2.3 was at stopping flu infections, they ran experiments on mice. They sprayed the protein into the noses of mice and then injected them with a heavy doses of influenza, which normally would be fatal.

    But the protein provided 100 percent protection from death. It remains to be seen if HB1.6928.2.3 can prove its worth in human trials.

    “It would be nice to have a front-line drug if a new pandemic was about to happen,” Dr. Wilson said.

    5
    In Dr. Baker’s office are models of complex proteins. The human body makes roughly 20,000, each suited to a different task. Credit Evan McGlinn for The New York Times

    HB1.6928.2.3 is just one of a number of proteins that Dr. Baker and his colleagues have designed and tested. They’ve also made a molecule that blocks the toxin that causes botulism, and one that can detect tiny amounts of the opioid fentanyl. Yet another protein may help people who can’t tolerate gluten by cutting apart gluten molecules in food.

    Last week, Dr. Baker’s team presented one of its most ambitious projects: a protein shell that can carry genes.

    The researchers designed proteins that assemble themselves like Legos, snapping together into a hollow sphere. In the process, they can also enclose genes and can carry that cargo safely for hours in the bloodstream of mice.

    These shells bear some striking resemblances to viruses, although they lack the molecular wherewithal to invade cells. “We sometimes call them not-a-viruses,” Dr. Baker said.

    A number of researchers are experimenting with viruses as a means for delivering genes through the body. These genes can reverse hereditary disorders; in other experiments, they show promise as a way to reprogram immune cells to fight cancer.

    But as the product of billions of years of evolution, viruses often don’t perform well as gene mules. “If we build a delivery system from the ground up, it should work better,” Dr. Baker said.

    Gary Nabel, chief scientific officer at Sanofi, said that the new research may lead to the invention of molecules we can’t yet imagine. “It’s a new territory, because you’re not modeling existing proteins,” he said.

    For now, Dr. Baker and his colleagues can only make short-chained proteins. That’s due in part to the cost involved in making pieces of DNA to encode proteins.

    But that technology is improving so quickly that the team is now testing longer, bigger proteins that might do more complex jobs — among them fighting cancer.

    In cancer immunotherapy, the immune system recognizes cancer cells by the distinctive proteins on their surface. The immune system relies on antibodies that can recognize only a single protein.

    Dr. Baker wants to design proteins that trigger a response only after they lock onto several kinds of proteins on the surface of cancer cells at once. He suspects these molecules will be better able to recognize cancer cells while leaving healthy ones alone.

    Essentially, he said, “we’re designing molecules that can do simple logic calculations.” Indeed, he hopes eventually to make molecular machines.

    Our cells generate fuel with one such engine, a gigantic protein called ATP synthase, which acts like a kind of molecular waterwheel. As positively charged protons pour through a ring of amino acids, it spins a hundred times a second. ATP synthase harnesses that energy to build a fuel molecule called ATP.

    It should be possible to build other such complex molecular machines as scientists learn more about how big proteins take shape, Dr. Baker said.

    “There’s a lot of things that nature has come up with just by randomly bumbling around,” he said. “As we understand more and more of the basic principles, we ought to be able to do far better.”

    See the full article here .

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  • richardmitnick 9:51 am on October 2, 2017 Permalink | Reply
    Tags: , , , , , The Baker Lab U Washington   

    From U Washington Medicine: “Mini-protein rapid design opens way to new class of drugs” 

    U Washington

    University of Washington

    September 27, 2017
    Leila Gray
    206.685.0381
    leilag@uw.edu

    Scientists at the Institute for Protein Design have created a way to generate thousands of different mini-protein binders as possible drug candidates. The proteins can be custom tailored to specific therapeutic targets. Recently, a set of these proteins were successfully tested in mice against the flu, and another group of these binders was able to protect brain cells against the botulism neurotoxin.

    1
    Artist impression of designed mini-protein binders targeting Influenza hemagglutinin to effectively bind and neutralize the virus. Cognition Studio, Daniel-Adriano Silva, Lance Stewart

    These computer-designed proteins, which did not previously exist in nature, combine the stability and bioavailability of small molecule drugs with the specificity and potency of larger biologics. They would not require refrigeration, and they likely would be simple for patients to take.

    “These mini-protein binders have the potential of becoming a new class of drugs that bridge the gap between small molecule drugs and biologics. Like monoclonal antibodies, they can be designed to bind to targets with high selectivity, but they are more stable and easier to produce and to administer,” said David Baker, who led the multi-institutional research project. He is a professor of biochemistry at the University of Washington School of Medicine and director of the UW Institute for Protein Design.

    Dr. David Baker, Baker Lab, U Washington

    Baker and his colleagues report their findings in article published online Sept. 27 by the journal Nature.

    Aaron Chevalier, Daniel-Adriano Silva and Gabriel J. Rocklin were the lead authors and were all senior fellows at the UW Institute for Protein Design at the time of the project.

    The method used a computer platform, called Rosetta, developed by Baker and colleagues at the University of Washington. They designed thousands of short proteins, about 40 amino acids in length, that the Rosetta program predicted would bind tightly to the molecular target.

    Rosetta@home project, a project running on BOINC software from UC Berkeley


    Rosetta@home BOINC project

    BOINC

    My BOINC

    Because of their small size, these short proteins tend to be extremely stable. They can be stored without refrigeration. They also are more easily administered than large protein drugs, such as monoclonal antibodies.

    Previously, such short, protein-binder drugs were typically re-engineered versions of naturally occurring proteins. These, however, tended not to be significantly better than monoclonal antibodies.

    Because these mini-proteins binders are original designs, they can be tailored to fit their targets much more tightly and are simpler to modify and refine.

    In this study, the researchers sought to design two sets of these proteins: one set that would prevent the influenza virus from invading cells and another that would bind to and neutralize a deadly nerve toxin from botulism. This toxin is considered a potential bioweapon.

    The computer modeling identified the amino-acid sequences of thousands of short proteins that would fit into and bind to the influenza and botulinum targets. The researchers created short pieces of DNA that coded each of these proteins, grew the proteins in yeast cells, and then looked at how tightly they bound to their targets. The targets were Influenza H1 hemagglutinin and botulinum neurotoxin B.

    All told, the method allowed them to design and test 22,660 proteins in just a few months. More than than two-thousand of them bound to their targets with high affinity.

    Evaluation of the best candidates found that the anti-influenza proteins neutralized viruses in cell culture and other designed proteins prevented the botulinum toxin from entering brain cells.

    A nasal spray containing one of the custom-designed proteins completely protected mice from the flu if administered before or as much as 72 hours after exposure.. The protection that the treatment provides equaled or surpassed that seen with antibodies, the researchers report.

    Testing of a subset of the proteins showed that they were extremely stable and, unlike antibodies, did not become inactivated by high temperatures. The small proteins also triggered little or no immune response, a problem that often renders larger protein drugs ineffective.

    Funding for the study came from Life Sciences Discovery Fund Launch grant (9598385), Doctorado en Ciencias Bioquiacutemicas UNAM (R56AI117675), Molecular Basis of Viral Pathogenesis Training Grant (T32AI007354-26A1), Investigator in the Pathogenesis of Infectious Disease award from the Burroughs Wellcome Fund and NIH (1R01NS080833), CoMotion Mary Gates Innovation Fellow; Shenzhen Science and Technology Innovation Committee (JCYJ20170413173837121), Hong Kong Research Grant Council (C6009-15G and AoE/P-705/16), PAPIIT UNAM (IN220516), CONACyT (254514) and Facultad de Medicina UNAM (AI091823, AI123920,AI125704), NIAID grant (1R41AI122431) (1R21AI119258), and Life Sciences Discovery Fund grant (20040757).

    See the full article here .

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    u-washington-campus
    The University of Washington is one of the world’s preeminent public universities. Our impact on individuals, on our region, and on the world is profound — whether we are launching young people into a boundless future or confronting the grand challenges of our time through undaunted research and scholarship. Ranked number 10 in the world in Shanghai Jiao Tong University rankings and educating more than 54,000 students annually, our students and faculty work together to turn ideas into impact and in the process transform lives and our world. For more about our impact on the world, every day.

    So what defines us — the students, faculty and community members at the University of Washington? Above all, it’s our belief in possibility and our unshakable optimism. It’s a connection to others, both near and far. It’s a hunger that pushes us to tackle challenges and pursue progress. It’s the conviction that together we can create a world of good. Join us on the journey.

     
  • richardmitnick 2:07 pm on June 16, 2017 Permalink | Reply
    Tags: , , , , The Baker Lab U Washington   

    From Science: “Designer protein halts flu” 

    AAAS
    Science Magazine

    June 12, 2017
    Robert Service

    1

    A designer protein (brown and orange) fits snugly on top of the influenza virus’s hemagglutinin protein (green), which helps the virus latch onto and infect cells.
    Eva-Maria Strauch

    There’s a new weapon taking shape in the war on flu, one of the globe’s most dangerous infectious diseases. Scientists have created a designer protein that stops the influenza virus from infecting cells in culture and protects mice from getting sick after being exposed to a heavy dose of the virus. It can also be used as a sensitive diagnostic. And although it isn’t ready as a treatment itself, the protein may point the way to future flu drugs, scientists say.

    “It’s impressive,” says James Crowe, an immunologist at Vanderbilt University in Nashville, who was not involved in the study. But because it hasn’t yet been tested in humans, “it [still] has a long way to go,” he says.

    Influenza severely sickens 3–5 million people each year, and it kills between 250,000 and 500,000, mostly the elderly and people with weakened immune systems. Every year, public health officials survey the three flu subtypes circulating in humans and design a vaccine for the next winter season that covers them all. But those vaccines are far from perfect: They don’t always exactly match the viruses actually going around, and in some people, the shots fail to trigger a vigorous immune response.

    Drugs are another line of defense. Most focus on the proteins on the virus’s outer coat, neuraminidase and hemagglutinin (HA). Some drugs that block neuraminidase, which helps the virus escape already infected cells, are starting to bump up against viral resistance. HA is scientists’ next target. The mushroom-shaped protein specializes in infecting cells, first by binding a trio of sites on its head to three separate sugar molecules on the surface of targeted cells. Once the virus latches on, parts of HA’s stem act as a grappling hook to pull the virus in close, allowing it to fuse with the cell membrane and release its contents inside.

    In 2011, researchers led by David Baker, a computational biologist at the University of Washington in Seattle, created a designer protein that binds HA’s stem, which prevented viral infection in cell cultures.

    Dr. David Baker, Baker Lab, U Washington

    But because the stem is often shrouded by additional protein, it can be hard for drugs to reach it.

    Now, Baker’s team has designed proteins to target HA’s more exposed head group. They started by analyzing x-ray crystal structures that show in atomic detail how flu-binding antibodies in people grab on to the three sugar-binding sites on HA’s head. They copied a small portion of the antibody that wedges itself into one of these binding sites. They then used protein design software called Rosetta to triple that head-binding section, creating a three-part, triangular protein, which the computer calculated would fit like a cap over the top of HA’s head group.

    Rosetta@home project, a project running on BOINC software from UC Berkeley

    My BOINC

    Next, they synthesized a gene for making the protein and inserted it into bacteria, which cranked out copies for them to test.

    In the test, Baker’s team immobilized copies of the protein on a paperlike material called nitrocellulose. They then exposed it to different strains of the virus, which it grabbed and held. “We call it flu glue, because it doesn’t let go,” Baker says. In other experiments, the protein blocked the virus from infecting cells in culture, and it even prevented mice from getting sick when administered either 1 day before or after viral exposure, they report today in Nature Biotechnology.

    Despite these early successes, Baker and Crowe caution that the newly designed protein isn’t likely to become a medicine itself. For starters, Baker says, the protein doesn’t bind all flu strains that commonly infect humans. That means a future drug may require either a cocktail of HA head group binding proteins or work in combination with stem-binding versions. Second, the safety of designer proteins will have to be studied carefully, Crowe says, because they are markedly different than natural HA-binding antibodies. “The further you get away from a natural antibody, the less you can predict what will happen,” Crowe says.

    But down the road, Baker says, the new designer protein could serve as the basis for a cheap diagnostic—akin to a pregnancy test—for detecting flu and possibly even medicines able to knock it out.

    See the full article here .

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