Directed evolution involves several key steps: 1. Mutation: Introducing random mutations into the gene encoding the enzyme. This can be done using techniques such as error-prone PCR or DNA shuffling. 2. Expression: Expressing the mutated genes in a suitable host organism, usually a bacterium like E. coli. 3. Screening/Selection: Identifying variants with the desired catalytic properties. This step often involves high-throughput screening methods to rapidly evaluate thousands of mutants. 4. Iteration: Repeating the process with the best-performing variants to further refine their properties.
