Directed evolution converts subtilisin E into a functional equivalent of thermitase.

By Huimin Zhao, Frances H. Arnold. 1999

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Phage-assisted continuous evolution of proteases with altered substrate specificity (2017, methodology): “In order to evolve high levels of activity on the final target substrate from these weakly active mutants, we applied three strategies to increase selection stringency on all three trajectories: (1) express a lower concentration of the PA-RNAP substrate by using a weaker constitutive promoter (proA ”
Hyperthermophilic Enzymes: Sources, Uses, and Molecular Mechanisms for Thermostability (2001, unknown): “engineered mesophilic enzymes [116, 374]) also suggests that”
Inferring Stabilizing Mutations from Protein Phylogenies: Application to Influenza Hemagglutinin (2009, unknown): “subtilisin [111], 0.”
Iterative saturation mutagenesis (ISM) for rapid directed evolution of functional enzymes (2007, methodology): “is the most frequently used method, with three to eight cycles of epPCR being typical, as, for example, in the enhancement of thermostabilit”
How Protein Stability and New Functions Trade Off (2008, background): “However, the generality of stability-function tradeoffs with regard to new functions should be addressed in view of the fact that, regardless of their relevance to function, most mutations are destabilizing [30,35–37].”