YESS 2.0, a Tunable Platform for Enzyme Evolution, Yields Highly Active TEV Protease Variants
Carl A. Denard, Chelsea Paresi, Rasha M. Yaghi, Natalie McGinnis, Zachary Bennett, Yi Li, George Georgiou, Brent L. Iverson
Abstract
). In our analysis, eTEV specifically digests a fusion protein in 2 h at a low 1:200 enzyme to substrate ratio. Structural modeling indicates that the increase in catalytic efficiency of eTEV is likely due to an enhanced interaction between the catalytic Cys151 with the P1 substrate residue (Gln). Furthermore, the modeling showed that the ENLYFQS peptide substrate is buried to a larger extent in the active site of eTEV compared with WT TEV. The new eTEV variant is functionally the fastest TEV variant reported to date and could potentially improve efficiency in any TEV application.
Topics & Concepts
Enzyme kineticsProteaseCatalytic efficiencyProteasesSubstrate (aquarium)EnzymeProtein engineeringPeptideDirected evolutionSubstrate specificityComputational biologyActive siteYeastBiochemistryChemistryBiologyGeneEcologyMutantViral Infectious Diseases and Gene Expression in InsectsCRISPR and Genetic EngineeringRNA and protein synthesis mechanisms