Microsecond acquisition of heterogeneous structure in the folding of a TIM barrel protein
Graduate School of Biomedical Sciences; Department of Biochemistry and Molecular Pharmacology
Medical Subject Headings
Anisotropy; Crystallography, X-Ray; Models, Molecular; *Protein Folding; Protein Structure, Secondary; Protein Structure, Tertiary; Spectrum Analysis; Thermodynamics; Time Factors; Triose-Phosphate Isomerase; Tryptophan Synthase
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.
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Citation: 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
DOI of Published Version
Proceedings of the National Academy of Sciences of the United States of America
Wu, Ying; Kondrashkina, Elena; Kayatekin, Can; Matthews, C. Robert; and Bilsel, Osman, "Microsecond acquisition of heterogeneous structure in the folding of a TIM barrel protein" (2008). GSBS Student Publications. 1385.