Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

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Record number 372287
Title Extensive formation of off-pathway species during folding of an alpha-beta parallel protein is due to docking of (non)native structure elements in unfolded molecules
Author(s) Nabuurs, S.M.; Westphal, A.H.; Mierlo, C.P.M. van
Source Journal of the American Chemical Society 130 (2008)50. - ISSN 0002-7863 - p. 16914 - 16920.
DOI https://doi.org/10.1021/ja803841n
Department(s) Biochemistry
VLAG
Publication type Refereed Article in a scientific journal
Publication year 2008
Keyword(s) azotobacter-vinelandii apoflavodoxin - nmr chemical-shifts - 8 m urea - secondary structure - denatured state - hydrogen-exchange - energy landscape - intermediate - topology - conformations
Abstract Detailed information about unfolded states is required to understand how proteins fold. Knowledge about folding intermediates formed subsequently is essential to get a grip on pathological aggregation phenomena. During folding of apoflavodoxin, which adopts the widely prevalent ¿¿ß parallel topology, most molecules fold via an off-pathway folding intermediate with helical properties. To better understand why this species is formed, guanidine hydrochloride-unfolded apoflavodoxin is characterized at the residue level using heteronuclear NMR spectroscopy. In 6.0 M denaturant, the protein behaves as a random coil. In contrast, at 3.4 M denaturant, secondary shifts and 1H¿15N relaxation rates report four transiently ordered regions in unfolded apoflavodoxin. These regions have restricted flexibility on the (sub)nanosecond time scale. Secondary shifts show that three of these regions form ¿-helices, which are populated about 10% of the time, as confirmed by far-UV CD data. One region of unfolded apoflavodoxin adopts non-native structure. Of the ¿-helices observed, two are present in native apoflavodoxin as well. A substantial part of the third helix becomes ß-strand while forming native protein. Chemical shift changes due to amino acid residue replacement show that the latter ¿-helix has hydrophobic interactions with all other ordered regions in unfolded apoflavodoxin. Remarkably, these ordered segments dock non-natively, which causes strong competition with on-pathway folding. Thus, rather than directing productive folding, conformational preorganization in the unfolded state of an ¿¿ß parallel-type protein promotes off-pathway species formation.
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