Synthesis and purification of site-specifically modified T7 RNA polymerase for bioorthogonal conjugation applications

Abstract

Aim. To develop protocol for site-specific introduction of bioorthogonal reaction groups into T7 RNAP while preserving activity of the enzyme. Conclusions. For both systems, the optimized protocols were developed, which allow successful incorporation of the bioorthogonal groups with near-quantitative modification efficacy and practical yields. Due to a greater flexibility of pAzF system for tether group positioning it will be used for immobilization studies.