Purification of AGO2 protein and investigation of micro RNA interactions

In this study, immunoaffinity purification of Argonaute 2 (AGO2) protein, known as RNA interference effector protein, preparation of nanoprotein particles having AGO2 protein and determination of micro RNA (miRNA) binding activities of these prepared nanoprotein particles have been aimed. In the first step, poly(HEMA-co-anti-AGO2) cryogel has been synthesized using photosensitive ruthenium- based amino acid monomer, bis(2-2ʹ-bipyridyl)(MATrp)2-ruthenium (II) according to ANADOLUCA method and characterized by FT-IR, SEM and water retention capacity test. Then, AGO2 protein has been purified via immunoaffinity chromatography from bovine liver cytoplasmic extract in a single step using this prepared cryogel. SDS- PAGE method has been used to determine the molecular weight and purity of this purified sample. The EMSA method has been used to exact determination of the presence of AGO2 protein in the purified sample. At the last stage, AGO2-based nano block polymers have been prepared using photosensitive microemulsion polymerization method. This prepared particles have been characterized by SEM and Zeta-Sizer and the binding affinities to commercial miR-335 have been determined. As a result, it has been determined that miR-335 has bound to AGO2 particles with a high affinity.