Abstract
By exploring the folding pathways of the B1 domain of protein L with a series of equilibrium and rapid kinetic experiments, we have found its unfolded state to be more complex than suggested by two-state folding models. Using an ultrarapid mixer to initiate protein folding within ~2-4 microseconds, we observe folding kinetics by intrinsic tryptophan fluorescence and fluorescence resonance energy transfer. We detect at least two processes faster than 100 μs that would be hidden within the burst phase of a stopped-flow instrument measuring tryptophan fluorescence. Previously reported measurements of slow intramolecular diffusion are commensurate with the slower of the two observed fast phases. These results suggest that a multidimensional energy landscape is necessary to describe the folding of protein L, and that the dynamics of the unfolded state is dominated by multiple small energy barriers.
| Original language | English |
|---|---|
| Pages (from-to) | 388-395 |
| Number of pages | 8 |
| Journal | HFSP Journal |
| Volume | 2 |
| Issue number | 6 |
| DOIs | |
| State | Published - Dec 2008 |
| Externally published | Yes |
Bibliographical note
Funding Information:This work was partially supported by funding from NSF FIBR Grant 0623664. The research of Lisa Lapidus, Ph.D. is supported in part by a Career Award at the Scientific Interface from the Burroughs Wellcome Fund. Work at Lawrence Livermore National Laboratory was performed under the auspices of the U.S. Department of Energy under Contract DE-AC52-07NA27344 with funding from the LDRD program. This work was partially supported by funding from NSF FIBR Grant 0623664 administered by the Center for Biophotonics, an NSF Science and Technology Center, managed by the University of California, Davis, under Cooperative Agreement PHY 0120999. L.J.L., W.J.W., S.A.W., and O.B. designed the experiments. S.A.W., S.J.D., V.R.S., Y.C., M.K., S.Y., and L.J.L. took the data. S.A.W., S.Y., and O.B. designed and fabricated the mixing chips. T.B., M.J., and M.K. mutated, expressed, and labeled the proteins. L.J.L., S.A.W., and W.J.W analyzed the data and wrote the paper.
Funding
This work was partially supported by funding from NSF FIBR Grant 0623664. The research of Lisa Lapidus, Ph.D. is supported in part by a Career Award at the Scientific Interface from the Burroughs Wellcome Fund. Work at Lawrence Livermore National Laboratory was performed under the auspices of the U.S. Department of Energy under Contract DE-AC52-07NA27344 with funding from the LDRD program. This work was partially supported by funding from NSF FIBR Grant 0623664 administered by the Center for Biophotonics, an NSF Science and Technology Center, managed by the University of California, Davis, under Cooperative Agreement PHY 0120999. L.J.L., W.J.W., S.A.W., and O.B. designed the experiments. S.A.W., S.J.D., V.R.S., Y.C., M.K., S.Y., and L.J.L. took the data. S.A.W., S.Y., and O.B. designed and fabricated the mixing chips. T.B., M.J., and M.K. mutated, expressed, and labeled the proteins. L.J.L., S.A.W., and W.J.W analyzed the data and wrote the paper.
| Funders | Funder number |
|---|---|
| Center for Biophotonics | |
| NSF Science and Technology Center | |
| National Science Foundation | 0623664 |
| U.S. Department of Energy | DE-AC52-07NA27344 |
| Burroughs Wellcome Fund | |
| Laboratory Directed Research and Development | |
| University of California, Davis | PHY 0120999 |