04/19/18 -9:45 AM to 11:00 AM
Kolthoff Lectureship #2: Professor Carlos J. Bustamante
Izaak M. Kolthoff Lectureship in Chemistry
"The Ribosome Modulates Nascent Protein Folding and Nascent Protein Folding Can Modulate the Ribosome Activity"
Proteins are synthesized by the ribosome and must attain their native structure to become functionally active. A great disparity exists between the rate of folding and the rate of synthesis of a polypeptide. The question then arises: what prevents the nascent polypeptide to misfold and become kinetically trapped before all of the information encoded in the sequence has emerged from the ribosome tunnel? Although it is commonly assumed that the ribosome affects the folding process, this idea has been difficult to demonstrate. We have developed an experimental system to investigate the folding of single ribosome-bound stalled nascent polypeptides of T4 lysozyme using optical tweezers. We show that the ribosome slows the formation of stable tertiary interactions and the attainment of the native state relative to the free protein. Incomplete T4 lysozyme polypeptides misfold and aggregate when free in solution, but they remain folding-competent near the ribosomal surface. Altogether, our results suggest that the ribosome not only decodes the genetic information and synthesizes polypeptides, but also promotes efficient de novo attainment of the native state, preventing the premature kinetic trapping of earlier fragments.
Protein synthesis rates can affect gene expression and the folding and activity of the translation product. Interactions between the nascent polypeptide and the ribosome exit tunnel represent one mode of regulating synthesis rates. The SecM protein arrests its own translation, and release of arrest at the translocon has been proposed to occur by mechanical force. Using optical tweezers, we demonstrate that arrest of SecM-stalled ribosomes can indeed be rescued by force alone and that the force needed to release stalling can be generated in vivo by a nascent chain folding near the ribosome tunnel exit. We formulate a kinetic model describing how a protein can regulate its own synthesis by the force generated during folding, tuning ribosome activity to structure acquisition by a nascent polypeptide.
Professor Bustamante is a professor of chemistry, physics, and molecular and cell biology, and the Raymond and Beverly Sackler Professor in Biophysics at the University of California, where he has been a faculty member and researcher since 1998. He received his bachelor’s degree from the Universidad Peruana Cayetano Heredia in Lima, Peru, his master's degree in biochemistry from the Universidad Nacional Mayor de San Marcos in Lima, and his doctorate in biophysics from University of California, Berkeley, where he studied with Ignacio Tinoco Jr. As a post-doctoral fellow at the Lawrence Berkeley National Laboratory, Bustamante studied with Marcos Maestre. Before moving to Berkeley, he was a professor at the University of Mexico, and professor of chemistry and member of the Insitute of Molecular Biology at the University of Oregon. He also was a Howard Hughes Medical Institute investigator. He is a Fellow of the American Physical Society, Fellow of the National Academy of Sciences, and Howard Hughes Medical Institute Investigator.
Professor Bustamante’s laboratory specializes in single molecule biophysics, where his researchers can track and measure the activity of individual enzymes. By looking at each enzyme, they can parse out effects which are not resolvable in bulk experiments. The researchers use optical tweezers, magnetic tweezers, atomic force microscopy, single molecule fluorescence, fluorescence correlation spectroscopy, and super-resolution photo-activatable light microscopy (PALM). They are interested in studying how the cell converts chemical energy into mechanical work through highly specialized molecular machines. The generation, transduction, and regulation of force are key to many central processes in the cell. Many enzymes, such as polymerases, are motors which move along a cellular track, using chemical energy to take regulated steps and control synthesis. These techniques are used to understand:
- DNA packaging into bacteriophage Φ29 capsids;
- translation and protein folding;
- protein degradation by ClpXP;
- PALM-mitochondrial fission, and
- catalysis-enhanced enzyme diffusion.
His group is affiliated with the departments of Molecular and Cell Biology, Physics, and Chemistry as well as the Physical Biosciences Division at the Lawrence Berkeley National Laboratory.
Kolthoff Lectureship in Chemistry
Izaak Maurits Kolthoff was born on February 11, 1894, in Almelo, Holland. He died on March 4, 1993, in St. Paul, Minnesota. In 1911, he entered the University of Utrecht, Holland. He published his first paper on acid titrations in 1915. On the basis of his world-renowned reputation, he was invited to join the faculty of the University of Minnesota’s Department of Chemistry in 1927. By the time of his retirement from the University in 1962, he had published approximately 800 papers. He continued to publish approximately 150 more papers until his health failed. His research, covering approximately a dozen areas of chemistry, was recognized by many medals and memberships in learned societies throughout the world, including the National Academy of Sciences and the Nichols Medal of the American Chemical Society. Best known to the general public is his work on synthetic rubber. During World War II, the government established a comprehensive research program at major industrial companies and several universities, including Minnesota. Kolthoff quickly assembled a large research group and made major contributions to the program. Many of Kolthoff’s graduate students went on to successful careers in industry and academic life and, in turn, trained many more. In 1982, it was estimated that approximately 1,100 Ph.D. holders could trace their scientific roots to Kolthoff. When the American Chemical Society inaugurated an award for excellence in 1983, he was the first recipient.