Improved method for capturing proteins holds promise for biomedical research


August 16, 2011

Antibodies are the backbone of the immune system – capable of targeting proteins associated with infection and disease. They are also vital tools for biomedical research, the development of diagnostic tests and for new therapeutic remedies.

Producing antibodies suitable for research however, has often been a difficult, costly and laborious undertaking. Download Full Image

Now, John Chaput and his colleagues at the Biodesign Institute at Arizona State University have developed a new way of producing antibody-like binding agents and rapidly optimizing their affinity for their target proteins. Such capture reagents are vital for revealing the subtleties of protein function, and may pave the way for improved methods of detecting and treating a broad range of diseases.

The team’s results appear in today’s issue of the journal ChemBioChem.

Antibodies are Y-shaped structures, capable of binding in two or more places with specific target proteins. Synthetic antibodies are much simpler forms that attempt to mimic this behavior. As Chaput explains, creating affinity reagents with strong binding properties can be accomplished by combining two weak affinity segments on a synthetic scaffold. The resulting affinity reagent, if properly constructed, can amplify the binding properties of the individual segments by two or three orders of magnitude.

“This dramatic change in affinity has the ability to transform ordinary molecules into a high affinity synthetic antibody,” Chaput says. “Unfortunately, the chemistry used to make these reagents can be quite challenging and often requires a lot of trial-and-error.  With NIH funding, my group has reduced the complexity of this problem to simple chemistry that is user friendly and easily amenable to high throughput automation. Such technology is absolutely necessary if we want to compete with traditional monoclonal antibody technology. ”

Traditionally, antibodies for research have been extracted from animals induced to produce them in response to various protein antigens. While the technique has been invaluable to medical science, obtaining antibodies in this way is a cumbersome and costly endeavor. Instead, Chaput and his team produce synthetic antibodies that do not require cell culture, in vitro selection or the application of complex chemistry. They call their reagents DNA synbodies.

The new strategy – referred to as LINC (for Ligand Interaction by Nucleotide Conjugates) uses DNA as a programmable scaffold to determine the optimal distance needed to transform two weak affinity binding segments or ligands into a single high affinity protein capture reagent. The result is an artificial antibody, capable of binding to its antigen target with both high affinity and high specificity. The process is rapid and inexpensive. It also offers considerable flexibility, as the distance between the two ligand components bonded to the short, double-stranded DNA scaffold can be fine-tuned for optimum affinity.

In earlier work, the group identified ligand candidates by producing thousands of random sequence peptide chains – strings of amino acids, connected like pearls on a necklace. The peptide sequences were affixed to a glass microarray slide and screened against a target protein to pinpoint those that were capable of recognizing distinct protein binding sites. Two promising ligand candidates could then be combined to form a DNA synbody.

In the current study, the group instead makes use of pre-existing ligands with documented affinity for various disease-related proteins. The method involves the use of well-characterized ligands as building components for high quality DNA synbodies, eliminating the initial screening procedure and expanding the potential to tinker with the two-piece synbody in order to optimize affinity.

The peptides of choice for the study were those with high affinity for something called growth factor receptor bound protein 2 (Grb2). Grb2 has many cell-signaling functions and is an important focus of research due to its association with cellular pathways involved in tumor growth and metastasis.

By scouring the scientific literature, the group identified two peptides that recognize distinct sites on the surface of Grb2.  Chaput points out, “this is a nice example where a few hours in the library can save you weeks in the lab.”

The next step was to create an assortment of synbody constructs based on these peptides. To do this, one peptide was attached to the end of a short DNA strand, while the other peptide was attached to the complementary DNA strand further along its length (see figure 1).

The two peptide strands could be attached to the scaffold in either a forward or reverse direction and could be interchanged, with either occupying the terminal end of the first DNA strand. Further, the distance between peptide segments along the DNA strands could be adjusted to yield the best target affinity. 

Experiments examined binding affinity for peptide chains separated by 3, 6, 9, 12, 15 and 18 base pairs along the DNA strand, (a distance range of 1.0-6.1 nm).  Inspection revealed the best results for a synbody constructed of peptides separated by 12 base pairs at a distance of 4.1 nm, compared with the other 5 constructs.

The results for the best synbody in the study were impressive, demonstrating a binding affinity five- to ten-fold stronger than commercially available antibodies for Grb2, despite the synbody’s comparatively primitive architecture. In further tests, the synbody was shown to exhibit high specificity – isolating Grb2 from other proteins in a complex biological mixture and selectively binding with its target. 

The technique offers a new approach to producing high qualityaffinity reagents for disease research, diagnostic testing and the development of effective therapeutics.

Richard Harth

Science writer, Biodesign Institute at ASU

480-727-0378

White House to feature student entrepreneur, business instructor


August 16, 2011

The White House will honor an innovative student who started his own business and an instructor from the W. P. Carey School of Business on Aug. 18. The two will be recognized at a Washington event designed to highlight young entrepreneurs and their importance to America’s future.

“Arizona State University has placed a major emphasis on encouraging student entrepreneurship, and the W. P. Carey School of Business continues to expand its strengths in this area,” says the school’s dean, Robert Mittelstaedt. “We understand the importance of entrepreneurial skills to all future business leaders, and we’re proud to have our students and faculty applauded for their achievements.” Sidnee Peck Download Full Image

Sidnee Peck, director of entrepreneurial initiatives at the W. P. Carey School of Business, is expected to speak at the White House about the benefits of teaching entrepreneurship to college students. Peck heads up a popular introductory entrepreneurship class that shows students how to turn their passion – anything from music to law to sports – into a business enterprise. Beginning this spring, Peck will also teach new classes called Semester to Launch and The Venture Capital Experience, which go deeper into launching and raising capital for businesses.

“These classes are open to all students, regardless of whether they are business majors,” explains Peck, a lecturer at the W. P. Carey School. “We want to give young people a firsthand understanding of what it really takes to make a business work, so they’ll have an advantage in the real world. Already, we’ve had some excellent students who are receiving a lot of attention.”

Among those high-achieving W. P. Carey School of Business students are Zach Hamilton and Jeremy Ellens. Ellens, a management major with a concentration in entrepreneurship, was recently named a finalist for Entrepreneur magazine’s “College Entrepreneur of 2011” award. He helped create a differential-diagnosis iPhone app that lets veterinarians evaluate animals’ symptoms and narrow down what illnesses they may have.

Zach Hamilton, another management major with a concentration in entrepreneurship, will be recognized at the White House this week for creating and owning Devil Wash. The successful, environmentally friendly pressure-washing business recently got him named an “All Star Student Entrepreneur” by Forbes magazine.

“I’m really excited to meet the other student entrepreneurs coming to the White House this week, to see what they’ve done and how they did it,” says Hamilton. “I’ve never been to Washington before, so I’m planning to see the sights and enjoy this.”

The event is part of the “Champions of Change” series in which the White House highlights ordinary Americans doing extraordinary things in their communities. It is likely both Peck and Hamilton will meet Vice President Joe Biden while there.

In addition to Peck’s popular entrepreneurship classes for undergraduates, the W. P. Carey School of Business also offers a certificate in small business and entrepreneurship. At the graduate level, the school recently announced expanded offerings in the area of leadership and a required class for all full-time and executive MBA students in entrepreneurship.

Also in the area of entrepreneurship, the school is doing groundbreaking research on how existing organizations can learn to be more entrepreneurial, especially with regard to continuous improvement. Plus, more entrepreneurship faculty members have been hired, and the Spirit of Enterprise Center continues to be a resource for small businesses in the Phoenix area. The center helps hundreds of businesses each year, offering companies the chance to recruit and meet with top student talent, while also allowing students to get hands-on business experience.

For more information about the W. P. Carey School of Business, go to www.wpcarey.asu.edu.