A Genetically Encoded Redox-Active Nicotinamide Amino Acid.

Nicotinamide-containing cofactors play an essential role in many enzymes that catalyze two-electron redox reactions. However, it is difficult to engineer nicotinamide binding sites into proteins due to the extended nature of the cofactor-protein interface and the precise orientation of the nicotinamide moiety required for efficient electron transfer to or from the substrate. To address these challenges, we genetically encoded a noncanonical amino acid (ncAA) bearing a nicotinamide side chain in bacteria.

An Orthogonal T7 Replisome for Continuous Hypermutation and Accelerated Evolution in .

Systems that perform continuous hypermutation of designated genes without compromising the integrity of the host genome can dramatically accelerate the evolution of new or enhanced protein functions. We describe an orthogonal DNA replication system in based on the controlled expression of the replisome of bacteriophage T7. The system replicates circular plasmids that enable high transformation efficiencies and seamless integration into standard molecular biology workflows.