The unfolding kinetics of ubiquitin captured with single-molecule force-clamp techniques Export

@article{schlierf:04, abstract = {10.1073/pnas.0400033101 We use single-molecule force spectroscopy to study the kinetics of unfolding of the small protein ubiquitin. Upon a step increase in the stretching force, a ubiquitin polyprotein extends in discrete steps of 20.3 \^{A}± 0.9 nm marking each unfolding event. An average of the time course of these unfolding events was well described by a single exponential, which is a necessary condition for a memoryless Markovian process. Similar ensemble averages done at different forces showed that the unfolding rate was exponentially dependent on the stretching force. Stretching a ubiquitin polyprotein with a force that increased at a constant rate (force-ramp) directly measured the distribution of unfolding forces. This distribution was accurately reproduced by the simple kinetics of an all-or-none unfolding process. Our force-clamp experiments directly demonstrate that an ensemble average of ubiquitin unfolding events is well described by a two-state Markovian process that obeys the Arrhenius equation. However, at the single-molecule level, deviant behavior that is not well represented in the ensemble average is readily observed. Our experiments make an important addition to protein spectroscopy by demonstrating an unambiguous method of analysis of the kinetics of protein unfolding by a stretching force.}, address = {Department of Biological Sciences, Columbia University, New York, NY 10027, USA.}, author = {Schlierf, Michael and Li, Hongbin and Fernandez, Julio M.}, citeulike-article-id = {1840050}, citeulike-linkout-0 = {http://dx.doi.org/10.1073/pnas.0400033101}, citeulike-linkout-1 = {http://www.pnas.org/content/101/19/7299.abstract}, citeulike-linkout-2 = {http://www.pnas.org/content/101/19/7299.full.pdf}, citeulike-linkout-3 = {http://view.ncbi.nlm.nih.gov/pubmed/15123816}, citeulike-linkout-4 = {http://www.hubmed.org/display.cgi?uids=15123816}, day = {11}, doi = {10.1073/pnas.0400033101}, issn = {0027-8424}, journal = {Proceedings of the National Academy of Sciences of the United States of America}, keywords = {afm, proteins, ubiquitin, unfolding}, month = {May}, number = {19}, pages = {7299--7304}, posted-at = {2009-10-26 06:26:40}, priority = {2}, title = {The unfolding kinetics of ubiquitin captured with single-molecule force-clamp techniques}, url = {http://dx.doi.org/10.1073/pnas.0400033101}, volume = {101}, year = {2004} }

TY - JOUR ID - schlierf:04 L3 - citeulike-article-id:1840050 N2 - 10.1073/pnas.0400033101 We use single-molecule force spectroscopy to study the kinetics of unfolding of the small protein ubiquitin. Upon a step increase in the stretching force, a ubiquitin polyprotein extends in discrete steps of 20.3 ± 0.9 nm marking each unfolding event. An average of the time course of these unfolding events was well described by a single exponential, which is a necessary condition for a memoryless Markovian process. Similar ensemble averages done at different forces showed that the unfolding rate was exponentially dependent on the stretching force. Stretching a ubiquitin polyprotein with a force that increased at a constant rate (force-ramp) directly measured the distribution of unfolding forces. This distribution was accurately reproduced by the simple kinetics of an all-or-none unfolding process. Our force-clamp experiments directly demonstrate that an ensemble average of ubiquitin unfolding events is well described by a two-state Markovian process that obeys the Arrhenius equation. However, at the single-molecule level, deviant behavior that is not well represented in the ensemble average is readily observed. Our experiments make an important addition to protein spectroscopy by demonstrating an unambiguous method of analysis of the kinetics of protein unfolding by a stretching force. IS - 19 JF - Proceedings of the National Academy of Sciences of the United States of America SN - 0027-8424 AD - Department of Biological Sciences, Columbia University, New York, NY 10027, USA. EP - 7304 TI - The unfolding kinetics of ubiquitin captured with single-molecule force-clamp techniques VL - 101 SP - 7299 KW - afm KW - proteins KW - ubiquitin KW - unfolding AU - Schlierf, Michael AU - Li, Hongbin AU - Fernandez, Julio PY - 2004/05/11/ UR - http://dx.doi.org/10.1073/pnas.0400033101 ER -