Dear all Apologies for the off-topic question: We are studying an enzyme that is activated by Ca2+. We obtained the crystals of the substrate and Ca2+-free form and solved the structure at 2.7 A resolution. However, the active site electron density map was not clear, although other regions are clear. We would like to determine the substrate-complex with Ca2+, which will elucidate the active site structure at a higher resolution. Now we have a problem that the crystals of a mutant which can bind the substrate and Ca2+ always have cracks in soaking to the crystallization solution containing CaCl2. Co-crystallization does not work at this time. We estimate the structural change in Ca2+-binding is not so big because the isozyme structure did not change very much when binding Ca2+. An isozyme structure in complex with the substrate was determined by soaking Ca2+ (and the substrate).
How should we overcome this problem? Best regards ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Masaki UNNO Graduate School of Science and Engineering, Ibaraki University, Japan
