Self-Association of Phosphorylase Kinase under Molecular Crowding Conditions

摘要

Self-association of phos-phorylase kinase (PhK) has been studied using analytical ultra-centrifugation and dynamic light scattering under the conditions of molecular crowding arising from the presence of high concentrations of osmolyte. Sedimentation velocity analysis shows that in accordance with the predictions of molecular crowding theory, trimethylamine N-oxide (TMAO) greatly favours self-association of PhK induced by Mg~(2+) and Ca~(2+). On the contrary, proline suppresses this process, probably, due to its specific interaction with PhK. We have also established that α-crystallin, a protein possessing chaperone-like activity, counteracts the self-association of PhK under mo- lecular crowding conditions. Using dynamic light scattering we have shown that the increase in the light scattering intensity accompanying self-association of PhK is due to the formation of particles having hydrodynamic radius of hundreds of nanometers. The hydrodynamic radius of the start associates (seeds of association) was found to be approximately 80 nm. TMAO facilitates the formation of the associates of larger size whereas proline and α-crystallin suppress self-association of PhK.