Dmitry G. Vassylyev
University of Alabama at Birmingham, USA
Title: A universal, ultra-high affinity chromatography for a one-step, high-throughput purification of complex proteins
Biography
Biography: Dmitry G. Vassylyev
Abstract
Protein purification is an essential, primary step in numerous biological studies, including a rapidly emerging proteomics field, its structural counterpart and drug screening. Moreover, purification with high yield, purity and activity (HHH-purification) is compulsory for high-resolution structural analysis and industrial protein production. Apparently, a universal, one-step high-throughput approach for HHH-purification is a key, yet unachievable, factor of success for all these crucial studies/applications. We designed and implemented an original chromatographic technology based on the physiological, ultra-high affinity (Kd~10-14-10-17 M) small (~10/16 kDa) protein/protein complex, which allows for a one-step HHH-purification of the traditionally challenging (eukaryotic, membrane, multi-subunit, DNA/RNA-binding, toxic) proteins. The developed system is practically indefinitely reusable, suitable for industrial-scale manufacturing, has high capacity (up to 20mg tagged protein per ml beads) and efficiently operates under nearly identical (target-independent) basic conditions. Altogether, it provides major advantages over the existing commercial analogs and may form a solid high-throughput platform for both, purification and associated “affinity” techniques (pull-down, kinetic activity/binding assays, etc.) of protein characterization.
