Unraveling multistate unfolding of rabbit muscle creatine kinase.
GdmCl-induced unfolding of rabbit muscle creatine kinase, CK, has been studied by a variety of physico-chemical methods including near and far UV CD, SEC, intrinsic fluorescence (intensity, anisotropy and lifetime) as well as intensity and lifetime of bound ANS fluorescence. The formation of several stable unfolding intermediates, some of which were not observed previously, has been established. This was further confirmed by representation of fluorescence data in terms of "phase diagram", i.e. I(lambda1) versus I(lambda2) dependence, where I(lambda1) and I(lambda2) are fluorescence intensity values measured on wavelengths lambda(1) and lambda(2) under the different experimental conditions for a protein undergoing structural transformations. The unfolding behavior of CK was shown to be strongly affected by association of partially folded intermediates. A model of CK unfolding, which takes into account both structural perturbations and association of partially folded intermediates has been elaborated.
Kuznetsova
Irina M
IM
Institute of Cytology, Russian Academy of Sciences, 194064 St. Petersburg, Russia.
Stepanenko
Olga V
OV
Turoverov
Konstantin K
KK
Zhu
Li
L
Zhou
Jun Mei
JM
Fink
Anthony L
AL
Uversky
Vladimir N
VN
D000818:Animals; D016880:Anisotropy; D002850:Chromatography, Gel; D002942:Circular Dichroism; D003402:Creatine Kinase / Q000737:chemistry*; D005453:Fluorescence; D006863:Hydrogen-Ion Concentration; D008962:Models, Theoretical; D018482:Muscle, Skeletal / Q000201:enzymology*; D017510:Protein Folding; D011817:Rabbits; D013056:Spectrophotometry, Ultraviolet
D003402:Creatine Kinase