Structural changes of creatine kinase upon substrate binding

Small-angle x-ray scattering was used to investigate structural changes upon binding of individual substrates or a transition state analog complex (TSAC; Mg-ADP, creatine, and KNO3) to creatine kinase (CK) isoenzymes (dimeric muscle-type (M)-CK and octameric mitochondrial (MI)-CK) and monomeric arginine kinase (AK). Considerable changes in the shape and the size of the molecules occurred upon binding of Mg-nucleotide or TSAC, The radius of gyration of MI-CK was reduced from 55.6 Angstrom (free enzyme) to 48.9 Angstrom (enzyme plus Mg-ATP) and to 48.2 Angstrom (enzyme plus TSAC), M-CK showed similar changes from 28.0 Angstrom (free enzyme) to 25.6 Angstrom (enzyme plus Mg-ATP) and to 25.5 Angstrom (enzyme plus TSAC). Creatine alone did not lead to significant changes in the radii of gyration, nor did free ATP or ADP. AK also showed a change of the radius of gyration from 21.5 Angstrom (free enzyme) to 19.7 Angstrom (enzyme plus Mg-ATP), whereas with arginine alone only a minor change could be observed. The primary change in structure as seen with monomeric AK seems to be a Mg-nucleotide-induced domain movement relative to each other, whereas the effect of substrate may be of local order only. In CK, however, additional movements have to be involved.