The influence of intersubunit cross-linking on the conformational changes of creatine kinase during denaturation by guanidine hydrochloride.
Intersubunit cross-linked creatine kinase (CK) has been prepared with the cross-linking reagent dithiobis(succinimidyl propionate) (DTSP). Unfolding of cross-linked CK during denaturation by guanidine hydrochloride (GuHCl), as monitored by intrinsic fluorescence, circular dichroism and fluorescence of the hydrophobic probe, 1-aniline-naphthalene-8-sulfonate (ANS), occurs in two stages with increasing GuHCl concentration. The process is similar to that of the unmodified enzyme, but in the second stage, conformational changes of the cross-linked enzyme need higher concentration of GuHCl, suggesting that there is a stable intermediate during its unfolding transition and the intermediate is stabilized by intersubunit cross-linkage.
Zhu
L
L
National Laboratory of Biomacromolecules, Institute of Biophysics, Academia Sinica, Beijing, China.
Fan
Y X
YX
Zhou
J M
JM
D000817:Anilino Naphthalenesulfonates; D002942:Circular Dichroism; D003402:Creatine Kinase / Q000737:chemistry*; D003432:Cross-Linking Reagents; D019281:Dimerization; D005456:Fluorescent Dyes; D019791:Guanidine; D011487:Protein Conformation; D011489:Protein Denaturation; D013050:Spectrometry, Fluorescence; D013388:Succinimides; D013438:Sulfhydryl Compounds / Q000737:chemistry
D000817:Anilino Naphthalenesulfonates; D003432:Cross-Linking Reagents; D005456:Fluorescent Dyes; D013388:Succinimides; D013438:Sulfhydryl Compounds; C027132:1-anilino-8-naphthalenesulfonate; D003402:Creatine Kinase; C011240:dithiobis(succinimidylpropionate); D019791:Guanidine