"We've discovered an interesting biochemistry question about how protein molecules interact with membranes, which happens to be relevant to biology and disease," Assistant Professor Jeff Knight says of the work he and his students are doing in a lab in the Science Building. "I hope it will continue to build for the rest of my career. Or at least until we solve the problem of Type 2 Diabetes."

The enthusiasm of chemistry colleagues Knight and Associate Professor Hai Lin is palpable. Both lean in when computer simulations of protein interactions with cell membranes start popping up on Knight's laptop screen. They animatedly explain the complex interactions about to take place. The work being done by Knight and Lin, along with other pioneers around the world, contributes to the search for cures to disease by providing a better understanding of how proteins play a role in cellular reactions. And their dedication is contagious. Students working in Knight's and Lin's labs are getting world-class research experience thanks to the success these professors have had in attracting funding, as well as their tireless dedication to training the next generation of biochemical researchers and modelers.

"Ever since I joined the Lin lab I have been fascinated by computational chemistry. I can create, modify, and analyze the biological systems using the computer," says Nara Chon, a student in the Chemistry BS/MS program. "I went to Norway to learn more advanced computer technique. Honestly, I really enjoyed the fact that I needed to not only integrate all the computational knowledge and skills but also communicate with all these experts in various chemical fields from different countries." Chon is honing these same skills in the lab at CU Denver and feels her research background will make her a strong applicant when she goes on to pursue her PhD in Computational Chemistry, "This is the reason why I chose this school."

Research this advanced can be hard to communicate, but Knight and Lin stress that when it comes to living organisms it's pretty much all about the proteins. Composed of strings of amino acids in tremendous varieties of sequences and forms, protein molecules participate in nearly all aspects of life at the cellular level. Studying how proteins inside a cell interact with the cell's membrane, or outer skin, is one very important key to understanding protein function—or in the case of Type 2 Diabetes—malfunction. Understanding insulin exocytosis (the process by which the insulin hormone is released from a cell into the bloodstream) is crucial in understanding the disorder.

"Insulin is packaged inside a membrane. There's also a membrane surrounding the cells themselves," begins Knight's instruction on the most basic level of his research. "In order to get this package of insulin out of a source cell membranes have to fuse, and proteins are responsible for doing that." All of the steps that make insulin available to do work are regulated by separate proteins, and Knight's lab is interested in a protein that senses accumulations of calcium inside a cell to trigger membrane fusion. He explains, "In Type 2 Diabetes, these cells that produce insulin gradually stop working. The body needs more and more insulin in a person who has Type 2 Diabetes as the disease progresses, and eventually these cells are not able to produce it. We want to understand how secretion happens at the molecular level because it has impacts on that."

Knight stresses that beyond this specific effect, protein signals are responsible at the cellular level for interactions going on all over the body, and understanding how those processes are taking place could have wide-ranging applications for researchers looking at many types of proteins. A closely related protein to the specific calcium-sensor protein Knight's lab is studying now (synaptotagmin 7) works in the brain to make synaptic connections between neurons (synaptotagmin 1). With a Research Corporation for Science Advancement (RCSA) grant, the Lin and Knight labs did research on this related protein starting in 2013. Knight and his students performed experiments to identify the key amino acid residues contributing to electrostatic and hydrophobic interactions. Using modeling techniques, Lin and his students compared the structures of the two protein molecules then employed computer simulations to theoretically alter the molecules' shapes to see how these changes affect their docking geometries and preferences. Understanding these differing affinities could have implications in changing the interactions and treating malfunctions.

Knight says, "The bottom line is the one involved in insulin secretion binds much more strongly to the membrane. We want to understand how two molecules with very similar structure can interact with membranes with such different affinity."

Last year, Knight was awarded a grant from the National Institutes of Health (NIH) to study just that. In Knight's lab experiments are conducted that collect data, and in Lin's lab the data are modeled for greater understanding, but they stress that the relationship between their research isn't that strictly linear. Knight says, "There are certain predictions that we would make, as experimentalists, we might say ‘Hey, it looks like our protein is inserting deeper into the membrane than that other one. Does that agree with what you've simulated?' and the simulations could lead Lin's team to say ‘Hey, it looks like this part of the protein is actually more important than we thought.' And then we can go in and see what happens if we go in and mutate that part of the protein."

As is the case with all great collaboration, Knight and Lin's labs achieve things together that they never could apart, including two back-to-back publications last year in the journal Biochemistry. The two papers are titled, "Membrane Docking of the Synaptotagmin 7 C2A Domain: Electron Paramagnetic Resonance Measurements Show Contributions from Two Membrane Binding Loops" (Osterberg, J.R.; Chon, N.L.; Boo, A.; Maynard, F.A.; Lin, H.; Knight, J.D.) and "Membrane docking of synaptotagmin-7 C2A domain: 2. Computations reveal interplay between electrostatic and hydrophobic contributions." (Chon, N. L.; Osterberg, J. R.; Henderson, J.; Khan, H.; Reuter, N.; Knight, J. D.; Lin, H.).

Knight came to CU Denver in 2010 from a Postdoc position at the University of Colorado Boulder. Shortly after meeting Lin, who had been modeling various biochemical reactions in the CU Denver Chemistry department since 2005, both began to see the potential in working together to attract funding and advance both of their individual labs' research. Cooperation also allows Knight and Lin to involve and inspire a wide range of students with various interests in biochemistry and modeling in the research process.

"I've always been interested in biochemistry, especially in relation to human health, and I suppose my interest showed through since Dr. Knight approached me one day after class to talk about his research and about joining his lab. After getting an overview of the types of projects I would be working on I knew his lab would be a perfect fit for me," says Ryan Osterberg, a recent MS graduate of the Chemistry program who now works in the medical laboratory testing field. "In the future I hope to move more and more into the research aspects of medical testing, hopefully assisting in development of new and better methods of disease detection."

Favinn Maynard is currently preparing to apply to competitive Medical Scientist Training Programs (MD/PhD), hoping to concentrate in neuroscience. She says, "When I first joined the Knight lab I was just looking for any type of research experience. I have come to really enjoy the research we do, because I find it fascinating that even though our research is on the molecular level it is forming part of the foundation that major diabetes research is based on."

The research results Maynard, Osterberg, and Chon helped produce alongside Knight and Lin have already led to a better understanding of how these particular proteins contribute to insulin secretion in both the healthy and diabetic states, and could possibly lead to new options for diabetes treatment or diagnosis. More broadly, studies being done by Knight, Lin, and the students who assist in their research will shed light on whether changes in membrane lipids may contribute to cellular defects that underlie many diseases. The possibilities for impacts on both human health and student futures that result from this partnership are endless.

Adapted in part from an article by Danielle Zieg, University Communications, June 4, 2013.