An optical method for the detection of oxidative stress using protein-RNA interaction

The cytosolic 4Fe-4S protein aconitase can be converted under the influence of reactive oxygen species into an iron-regulatory protein (IRP1). Therefo re, the IRP1 level is considered as an indirect marker of oxidative stress. An experimental approach is presented here to detect the concentration of this marker protein by surface plasmon resonance. The optical method exploi ts the natural binding affinity of IRP1 to an iron-responsive element (IRE) which was in vitro transcribed with a linker sequence and subsequently imm obilized on a BIACORE sensor chip. The detection was found to be reproducib le and sensitive in the range 20-200 nM MP. Conditions of the binding proce ss, such as pH and thiol concentration, were characterized. Feasibility of the method to detect and quantify IRP1 in physiological media was demonstra ted.