Collaborations Are Everything
Of all the collaborations in which Associate Professor M.G. Finn has participated since coming to The Scripps Research Institute (TSRI), few have been more rewarding than a day he spent in his laboratory making molecules last summer.
That day, he was entertaining and being entertained by his eight-year-old son, Marc, to whom he was giving a hands-on demonstration of the magic of organic synthesis and discovery. Earlier in the week, Finn had set up a synthetic reaction so that it was all but complete. The last stepsimply mixing two solutions togetherhe had saved for his son.
"When I handed [the beakers] to him," says Finn, "I said, 'You are making something that has never existed on earth before.'"
Finn's son then mixed the ingredients and the solution changed color (no stranger to the education process, Finn had set up the reaction this way). This brought a smile to the child's face, who realized the color change meant he had created something new. And Finn smiled as well, pleased to see his son as excited as he was about making molecules.
The more advanced lesson, what these molecules do, would wait for another day. For the moment, both father and son shared the joy of basic discovery, and were satisfied with their collaboration.
The Mechanism is the Message
Indeed, collaboration has been a satisfying way of life for Finn, who trained originally as an inorganic chemist and spent the bulk of his career in organometallic chemistry before pursuing an interest in biology after he heard a lecture by TSRI President Richard Lerner in the mid-1990s. Hoping to learn more molecular biology, Finn asked Lerner after the lecture about the possibility of doing a sabbatical at TSRI.
"I knew no molecular biology, and I wanted to learn," says Finn.
Finn arrived at TSRI in 1996 to do a year-long sabbatical while he was still a professor at the University of Virginia. He worked with Professor Carlos Barbas and Lerner on catalytic antibodieslooking at metal additives to the aldolase antibody system, a class of antibodies that are capable of catalyzing aldol reactions.
After he returned to Virginia, he was offered a position at TSRI, including an invitation to join The Skaggs Institute for Chemical Biology. He accepted, returning the next year with a small number from his group and working temporarily out of a few fume hoods in the laboratory of TSRI Professor and 2001 Nobel laureate K. Barry Sharpless. By 1999, Finn had transitioned his entire laboratory to TSRI and occupied space in the CVN building, where part of his group remains today.
Another part of his group occupies some of the brand-new laboratory space devoted to the Center for Integrative Molecular Biosciences (CIMBio) at TSRI, of which Finn is a founding member. CIMBio is a new collaborative effort whose mission is to foster multidisciplinary studies of molecular "machines," with the aim of determining their structure, their mechanism of action, and their dynamic behavior in the context of living cells.
CIMBio suits Finn well, because as a chemist, he loves to synthesize molecules, especially those that might be useful catalytic engines, and to study their mechanisms and how they behave. "What they do and how they do what they do," says Finn. "That's what I love to think about."
And his broad research goals are to use the reactivity of these moleculesespecially metal-containing onesto drive reactions in chemistry and biology. CIMBio provides him with lots of opportunities to collaborate with biologists and conduct research that asks what his molecules do in living cells and organisms.
Mass Spectrometry as an Analytical Tool
Finn has many other collaborations as well, including some that he started shortly after he first arrived at TSRI. One of his first collaborations was with Gary Siuzdak, who directs the Center for Mass Spectrometry at TSRI. Finn and Siuzdak have been using mass spectrometry as an analytical tool for the high-throughput screening of catalytic compounds and for measuring the efficiency of enantioselective chemical catalysisreactions that produce either right- or left-handed enantiomers.
"We're interested in profiling the activities of new and known catalysts, screening them against a variety of potential substrates all at once, and getting a rough idea of what is good and what is not good," says Finn.
For instance, the scientists can react different chiral catalysts with different "mass-tagged" chiral substrates, making products that can be read according to their masses. The mass spectrometry then enables the scientists to determine the most efficient catalysts, and Finn and his colleagues are applying this technique to many different catalytic reactions and many broad classes of substrates, including alcohols, epoxides, ketones, aldehydes, and olefins.
The technique is quick, sensitive enough to detect very small amounts of material, and does not require the substrate to be altered (by adding a fluorophor, for instance).
Finn and Siuzdak are also working to couple this technique with another, which Siuzdak has pioneered, that uses laser desorption/ionization of small molecules on porous silicon. Small amounts of substrate can be chemically attached to the porous silicon, and Finn and his coworkers have discovered a unique set of cleavable linkers that allows selective detachment of these molecules during the ionizing laser pulsethe first step in analyzing their mass molecules.
Another Research Project is Born
Finn's work with Professor Sharpless dates back long before Sharpless loaned him laboratory space when Finn first arrived from Virginia. Finn received his Ph.D. degree from Sharpless's group in the mid-1980s, and he and Sharpless had stayed in touch since. When Finn arrived at TSRI in the late 1990s, their longstanding collaboration was reborn.
Sharpless was working on a new idea called "click chemistry." Click chemistry, a modular protocol for organic synthesis that Sharpless developed, is a powerful and original approach to drug design. In its "in situ" variant, the target itself is recruited to play a decisive role in the synthesis of its own inhibitor in the last step.
The first target molecule that Sharpless and his colleagues worked on was acetylcholinesterase, a brain enzyme that breaks down acetylcholine, the neurotransmitter that propagates nerve signals. Inhibitors of acetylcholinesterase are used to treat the dementia associated with Alzheimer's disease, increasing the amount of acetylcholine in the brain, in turn enhancing brain activity.
Finn's student Warren Lewis, a Ph.D. candidate in TSRI's Kellogg School of Science and Technology, sorted out the kinetics of the system with the help of Sharpless and Finn's collaborator Palmer Taylor at the University of California, San Diego.
Finn notes that the mass spectrometry technique developed by his and Siuzdak's laboratories was instrumental in identifying the "hit" in the project. It enabled them to pick out the tiny amount of inhibitor that was synthesized with the help of the acetylcholinesterase enzyme. This success was reported last year by Lewis, Finn, Taylor, Sharpless, and several others in an article in the journal Angewandte Chemie.
The success of the acetylcholinesterase work also gave Sharpless and Finn the opportunity to join a program project grant directed by TSRI Professor Arthur Olson. The project seeks to establish a drug design cycle aimed at developing, testing, and refining novel approaches to making specific inhibitors that will hit resistant mutants of HIV protease.
"That enzyme," says Finn, "should, in principle, be amenable to the same kind of [click chemistry] strategy as acetylcholinesterase."
The strategy, he explains, is best applied where protein-protein interfaces exist. Such regions often have multiple potential binding sites for small moleculesas in the case of the HIV protease, a dimer formed by two identical protease monomers. Acetylcholinesterase itself is not a dimer but has two known binding regions adjacent to each other.
TSRI Assistant Professor Valery Fokin of the Sharpless lab is directing the synthesis of component molecules that will be used as building blocks for designing inhibitors. Other collaborators in the program project will provide mutant and wild-type protease, and the Sharpless, Fokin and Finn laboratories will attempt to hit them with these diverse blocks, as they did before with acetylcholinesterase, and fish out those combined molecules that the protease enzyme itself assembles.
"We want to let the enzyme teach us what inhibitors [it prefers]," says Finn. "Those, in general, should be the better inhibitors."
The Finn and Sharpless laboratories are also using click chemistry to develop new materials, and they have chosen to make adhesives first. They quickly found a metal adhesive that is far stronger than the glues currently sold for the purpose.
And Finn, chemist at heart that he is, has also continued to develop new synthetic methods and apply them to the biological targets of his collaborators. Finn's group recently published a new method to make a new class of pharmacophores by bringing together urea compounds with guanidine-like compounds. This has allowed them to create "tunable" electrophiles that react with a wide range of nucleophiles.
A Collaboration Built Upon Structures
Some time in late 1999, Finn wandered down to the laboratory of TSRI Professor John Johnson, whom he had recently met. As Finn describes it, the two of them then spent a "golden afternoon" in Johnson's laboratory looking at the viral structures that Johnson had been working on. Among these structures was the one of cowpea mosaic virus, an icosahedral RNA virus that infects the plant that produces black-eyed peas.
"I learned from Jack that you could make these [viruses] in gram quantities," says Finn. The structures of the virus particles were known from Johnson's work, and Finn realized as he was fiddling with some of the three-dimensional models that afternoon that they could be just the building blocks he was looking for. Finn saw these virions as "supramolecular" chemical reagents that can be manipulated to display a number of interesting molecules by attaching other chemicals to the side chains of the viral component proteins.
"At that moment, I proposed to Jack that we collaborate," says Finn.
And collaborate they did. As a member of The Skaggs Institute for Chemical Biology, Finn used funding that was provided to him by The Skaggs Institute for Research to get the project started. Johnson and TSRI Assistant Professor Tianwei Lin helped Finn's group get a handle on the molecular biology of the viruses, showing him how to manipulate their protein sequences and to express and purify them, and Finn contributed his organic synthetic experience to Johnson's virions, designing ways to attach new molecules to their surfaces.
The molecular biology of the virions can be tweaked so as to provide different kinds of "hooks" onto which different chemicals can be attached. The attachments are made after the particles are harvested, intact, from infected plants, and these particles are so sturdy that the chemistry can be done over a wide range of pH levels, temperatures, and organic solvent concentrations.
These attachments are made via lysine or cysteine side chains on the subunits of the proteins that come together to make the viral shell. Since there are multiple protein subunits and potentially multiple exposed lysine or cysteine side chains, multiple copies of the added chemical can be attached.
In so doing, it is possible to produce materials with a number of different properties and a variety of potential uses. The fertile ground created by this intersection of chemistry and biology was recognized in 2001 by the David and Lucile Packard Foundation, which made Finn and Johnson the recipients of its Interdisciplinary Science Program award in a national competition.
Finn's collaboration with Sharpless also allowed him to pioneer click chemistry as a method for making attachments to biological molecules, using viruses as the test case. This has been picked up by a number of laboratories, including that of Cell Biology and Chemistry Associate Professor Ben Cravatt, for making bonds in and around cells.
Drug Delivery and Materials Design
One of the most obvious uses of virus particles is biomedicalthe delivery of a drug to a particular tissue or cell type in the body, for instance. "Can you take this big particle, steer it to a particular cell type, and deliver a payload?" Finn asks.
The first step in this process, he says, is chemical sensing, or targeting a particle to a certain part of the body. For this targeting, Finn collaborates with Cell Biology Assistant Professor Marianne Manchester, who has an adjacent office and shares laboratory space in the CIMBio building. Manchester has specialized in following dye-decorated virus particles through whole animals and tailoring them genetically to find particular tissues. The goal of their collaboration is to turn the viruses into molecules that could report on disease states and perform drug delivery.
"I think we are getting pretty close," Finn says, adding that the next step is extending the technology to deliver a payload.
For the delivery, Finn works with Department of Cell Biology Chair Sandra Schmid to characterize the receptor-mediated endocytosis of virus particles with carbohydrates displayed on their surfaces. The hope is to be able to induce the cells to take up the virus particle selectively. One key to this is the fact that the virions are polyvalent, and multiple copies of some endocytosis "effector" molecule can be displayed on their surface in well-defined patterns and distances.
Finn is also working with Associate Professor Glen Nemerow in the Department of Immunology to try to design plant virus particles that mimic adenovirus, the virus that causes the common cold, which Nemerow studies. Adenovirus is already adept at getting into cells through a complicated binding and entry mechanism.
In a different twist, Finn and his laboratory are trying to use Johnson's virions to make new materials, and they have made progress in getting the virions to assemble themselves into supermolecular assemblies, aggregates, and other nanostructures by putting different molecules on the outside of the virions.
"This is the first step for us," says Finn, who adds that his goal is to be able to program the virions to make the emergent assemblies he desires.
To test this, graduate student Erica Strable, a Ph.D. candidate in TSRI's Kellogg School of Science and Technology who is a joint member of the Finn and Johnson laboratories and is funded by the La Jolla Interfaces in Science Program directed by TSRI Professor Libby Getzoff and UCSD Professor Jose Onuchic, chemically attached DNA oligonucleotides to the outside of the virus and created different pools of such oligovirus particles with complimentary bases. Depending on where the oligos were placed on the virus, Strable found that she could assemble the pieces into two-dimensional arrays or three-dimensional aggregates in a temperature-sensitive fashion (since high temperature melts DNA).
He also has been experimenting with assembling these particles by attaching antibodies to some and antigen to others, by using metal interactions and attaching sugars to some virions and carbohydrate binding proteins to others. All of these have potential uses in biology and nanotechnology, applications that Finn is currently exploring with his many collaborators
"The best part about being here," he says. "is all the fabulous people who are in my group and around the institute. I delight in being a mentor to the wonderful students and postdocs that I find here at Scripps, both in my group and in the graduate program."
"And," says Finn, "I'm having the time of my life."