THE EFFECTS OF FILTRATION ON PROTEIN NUCLEATION IN DIFFERENT GROWTH MEDIA

Filtration effects of turkey egg white lysozyme solution (TEWL) prior to subjecting it to crystallization conditions are investigated. Filte ring TEWL solution and crystallizing it in ungelled media significantl y decreased the number of conditions yielding crystals. This decrease depended on the membrane cut-off used for filtration. From this, the p ostulated factors aiding in nucleation are estimated to be 0.17 mu m i n diameter. The existence of these factors was verified by the procedu re of reversed filtration: filtered solutions passed through their inv erted filter membrane a second time lead to improved crystallization r esults. The effect of aging of the TEWL solution prior to subjecting i t to ungelled crystallization conditions was also verified. We did not find any time-dependent change in the size or the number of crystals per drop. Repeating the filtration experiments in agarose-gelled cryst allization media showed that the influence of filtration on the crysta llization outcome was significantly diminished. Far better crystalliza tion results were obtained compared to ungelled media. However, there is a certain aging effect linked to filtration in gelled media. Differ ent crystallization results were obtained depending on whether filtrat ion was performed before or after aging and subsequent crystallization . This suggests a secondary time-dependent effect.