The 3-D structures of CAP-Gly, which show the spatial arrangement of
atoms in the protein molecule, are different between the free state of
the protein and its bound state to the microtubule. These structures
reveal how the protein interacts with microtubules, predominantly
through its loop regions, which adopt specific conformations upon
However, static structures of CAP-Gly do not tell the whole story about the protein.
“Just as we are always moving our arms and legs about, proteins are
very dynamic. They do not stand still,” Polenova says. “These motions
are essential to their biological function, and NMR spectroscopy is the
only technique that can record such movements, with atomic resolution,
on a variety of time scales, from picoseconds to arbitrarily long time
scales — seconds, days, weeks — to help us understand the protein’s
function. We know from our prior studies that CAP-Gly is dynamic on
timescales from nano- to milliseconds, and this mobility is essential
for the protein’s ability to interact with microtubules and with
multiple other binding partners.”
The research, which has been ongoing since 2008 when the first data
sets were collected, required the development of new protocols for
preparing the samples, new NMR experiments to gather various information
on structure and dynamics, and new protocols for data analysis.
In the future, Polenova and her team envision using NMR in
combination with cryo-electron microscopy, in which samples are studied
at extremely low temperatures, typically below 200 degrees Fahrenheit,
to look at even more complex systems in a highly preserved form.
Polenova’s research team at UD included Si Yan, who received her
doctorate from the University in 2014, current doctoral student
Changmiao Guo, NMR spectroscopist Guangjin Hou and postdoctoral
researchers Huilan Zhang and Xingyu Lu. Williams, at Beckman Research
Institute, also was a co-author of the study.
The research was supported by the National Institutes of Health
through a grant from the National Institute of General Medical Sciences.
The National Science Foundation funded one of the NMR spectrometers used in the research.
Polenova and her students and colleagues published a second article,
on protein regulation of HIV activity, in the same issue of the Proceedings of the National Academy of Sciences. Read that UDaily article here.