Oral presentations-A comprehensive platform to investigate protein-metal ion interactions by affinity capillary electrophoresis (ACE)

摘要

Affinity Capillary Electrophoresis (ACE) provides an important enhancement to characterize molecular interactions. In exciting recent studies, the influence of various metal ions, including Li?, Na~+, Mg~(2+), Ca~(2+), Ba~(2+), Al~(3+), Ga~(3+), La~(3+), Pd~(2+), Ir~(3+), Ru~(3+), Rh~(3+), Pt~(2+), Pt~(4+), Os~(3+), Au~(3+), Au~+, Ag~+, Cu~(2+), Fe~(2+), Fe~(3+), Co~(2+), Ni~(2+), Cr~(3+), V~(3+), Mn~(2+), MoO_4~(2-) and SeO_3~(2-) was investigated by ACE, giving deep insight into the functional interactions between these species and biomolecules. The predominant role of ACE is in the early screening stage when binding and non-binding compounds are sorted out. The requirements for sample amount and purity are low, but high precision of binding information in reasonable short analysis times can be expected [1]. ACE can now be performed in *5 min including rinsing procedures. An excellent precision, corresponding to RSD % of 0.2–1.0 % was achieved. Long term stability and appropriate method transfers have also been established. The capillary manufacture batch, the type of temperature controlling tool, the purity of running buffer constituents and the quality of the ligands involved, including their stability, have been identified as main parameters for robustness. Further ACE key method development parameters include protein concentration, length of injected plug, applied voltage, and the choice of the regression method [2]. Now we not only provide a generic concept and experimental conditions for all relevant metal ions to be investigated, which could be easily enhanced to each and every further species, but we also provide reference values for characteristic interactions to a set of reference proteins. These concepts have already been successfully applied for a number of applications, namely Extracellular-signal Regulated Kinase (ERK), dehydrins (metal-ion storing plant proteins), potentially Ca~(2+) binding peptides and transferrin.