TY  - JOUR
AU  - Cerminara, Michele
AU  - Schöne, Antonie
AU  - Ritter, Ilona
AU  - Gabba, Matteo
AU  - Fitter, Jörg
TI  - Mapping Multiple Distances in a Multidomain Protein for the Identification of Folding Intermediates
JO  - Biophysical journal
VL  - 118
IS  - 3
SN  - 0006-3495
CY  - Bethesda, Md.
PB  - Soc.
M1  - FZJ-2020-00862
SP  - 688-697
PY  - 2020
AB  - The investigation and understanding of the folding mechanism of multidomain proteins is still a challenge in structural biology. The use of single-molecule Förster resonance energy transfer offers a unique tool to map conformational changes within the protein structure. Here, we present a study following denaturant-induced unfolding transitions of yeast phosphoglycerate kinase by mapping several inter- and intradomain distances of this two-domain protein, exhibiting a quite heterogeneous behavior. On the one hand, the development of the interdomain distance during the unfolding transition suggests a classical two-state unfolding behavior. On the other hand, the behavior of some intradomain distances indicates the formation of a compact and transient molten globule intermediate state. Furthermore, different intradomain distances measured within the same domain show pronounced differences in their unfolding behavior, underlining the fact that the choice of dye attachment positions within the polypeptide chain has a substantial impact on which unfolding properties are observed by single-molecule Förster resonance energy transfer measurements. Our results suggest that, to fully characterize the complex folding and unfolding mechanism of multidomain proteins, it is necessary to monitor multiple intra- and interdomain distances because a single reporter can lead to a misleading, partial, or oversimplified interpretation.
LB  - PUB:(DE-HGF)16
C6  - pmid:31916943
UR  - <Go to ISI:>//WOS:000511291400016
DO  - DOI:10.1016/j.bpj.2019.12.006
UR  - https://juser.fz-juelich.de/record/873619
ER  -