STRUCTURAL DYNAMICS OF CREATINE KINASE. Michael Forstner¹, M. Kriechbaum², P. Laggner² and T. Wallimann¹. ¹Institute for Cell Biology, ETH Zuerich, Switzerland, and ²Institute of Biophysics and X-Ray Structure Research, Austrian Acad. of Sciences, Graz, Austria

Small-angle X-ray (SAXS) and neutron scattering were used to investigate structural changes upon binding of individual substrates or a transition state analogue complex (TSAC), consisting of Mg-ADP, creatine and KNO₃ to creatine kinase isoenzymes (dimeric M-CK and octameric Mi-CK) and monomeric arginine kinase (AK). Considerable changes in the shape and the size of the molecules occured upon binding of Mg-ATP and TSAC, whereas neither creatine nor free nucleotide led to significant changes in CK structure. In Mi-CK, the radius of gyration was reduced from 55.6 (free enzyme) to 48.9 (enzyme + Mg-ATP) and to 48.2 (enzyme + TSAC). The experiments performed with M-CK showed similar changes from 28.0 (free enzyme) to 25.6 (enzyme+Mg-ATP, enzyme +TSAC). AK showed the same behaviour. Based on the X-ray structure coordinates of Mi-CK we have tried to model the nature of the structural transition found. The X-ray structure of Mi-CK in the presence of ATP is that of the "open" form as seen by comparison of the calculated and experimentally derived scattering curves. Homology modeling allowed for the derivation of models for M-CK and AK. The primary change in structure as seen with monomeric AK seems to be a magnesium-nucleotide induced domain movement relative to each other, whereas the effect of substrate may be of local order only. One highly mobile loop structure was identified, that might work as a lid to close the active site during catalysis. In CK, however, further movements must be involved in the large conformational change. In Mi-CK the appearance of compactness is concurrent with a change of the channel penetrating the octamer to a more cavity-like structure. Furthermore investigations on the time course of the structural transitions have been performed by time-resolved SAXS.