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Direct detection of the protein quaternary structure and denatured entity by small‐angle scattering: guanidine hydrochloride denaturation of chaperonin protein GroEL
Author(s) -
Seki Yasutaka,
Hiragi Yuzuru,
Ichimura Kaoru,
Soda Kunitsugu
Publication year - 2002
Publication title -
journal of applied crystallography
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.429
H-Index - 162
ISSN - 1600-5767
DOI - 10.1107/s002188980101473x
Subject(s) - small angle x ray scattering , guanidine , protein quaternary structure , radius of gyration , denaturation (fissile materials) , circular dichroism , chemistry , crystallography , random coil , protein folding , monomer , scattering , polymer , biochemistry , organic chemistry , nuclear chemistry , physics , protein subunit , optics , gene
A change in the higher‐order structure of an oligomeric protein is directly detectable by small‐angle scattering. A small‐angle X‐ray scattering (SAXS) study of the denaturation process of the chaperonin protein GroEL by guanidine hydrochloride (GdnHCl) showed that the disappearance of the quaternary structure can be monitored by using a Kratky plot of the scattered intensities, demonstrating the advantage of the SAXS method over other indirect methods, such as light scattering, circular dichroism (CD), fluorescence and sedimentation. The collapse of the quaternary structure was detected at a GdnHCl concentration of 0.8 M for a solution containing ADP (adenosine diphosphate)/Mg 2+ (2 m M )/K + . From pairwise plots of the change in forward scattering intensity J (0)/ C (weight‐average molecular weight) and the z ‐average (root mean square) radius of gyration against the GdnHCl concentration, the stability and nature of the denatured protein can be determined. The SAXS results suggest that the GroEL tetradecamer directly dissociates to the unfolded coil without going through a globular monomer state. The denatured ensemble is not a single unfolded monomer coil particle, but some mixture of entangled aggregates and a monomer of the coil molecules. Small‐angle scattering is a powerful method for the detection and study of changes in quaternary and higher‐order structures of oligomeric proteins.