Microsecond acquisition of heterogeneous structure in the folding of a TIM barrel protein
Graduate School of Biomedical Sciences; Department of Biochemistry and Molecular Pharmacology
Life Sciences | Medicine and Health Sciences
The earliest kinetic folding events for (betaalpha)(8) barrels reflect the appearance of off-pathway intermediates. Continuous-flow microchannel mixing methods interfaced to small-angle x-ray scattering (SAXS), circular dichroism (CD), time-resolved Forster resonant energy transfer (trFRET), and time-resolved fluorescence anisotropy (trFLAN) have been used to directly monitor global and specific dimensional properties of the partially folded state in the microsecond time range for a representative (betaalpha)(8) barrel protein. Within 150 micros, the alpha-subunit of Trp synthase (alphaTS) experiences a global collapse and the partial formation of secondary structure. The time resolution of the folding reaction was enhanced with trFRET and trFLAN to show that, within 30 micros, a distinct and autonomous partially collapsed structure has already formed in the N-terminal and central regions but not in the C-terminal region. A distance distribution analysis of the trFRET data confirmed the presence of a heterogeneous ensemble that persists for several hundreds of microseconds. Ready access to locally folded, stable substructures may be a hallmark of repeat-module proteins and the source of early kinetic traps in these very common motifs. Their folding free-energy landscapes should be elaborated to capture this source of frustration.
DOI of Published Version
Proc Natl Acad Sci U S A. 2008 Sep 9;105(36):13367-72. Epub 2008 Aug 29. Link to article on publisher's site
Proceedings of the National Academy of Sciences of the United States of America
Wu Y, Kondrashkina E, Kayatekin C, Matthews CR, Bilsel O. (2008). Microsecond acquisition of heterogeneous structure in the folding of a TIM barrel protein. GSBS Student Publications. https://doi.org/10.1073/pnas.0802788105. Retrieved from https://escholarship.umassmed.edu/gsbs_sp/1385