Ukierunkowana ewolucja enzymów pochodzenia mikrobiologicznego
1. Wprowadzenie. 2. Udoskonalanie enzymów metodami nierekombinacyjnymi. 3. Udoskonalanie enzymów metodami rekombinacyjnymi. 4. Podsumowanie
Abstract: Enzymes of microbial origin are extensively used in different industrial processes. However, very often these biocatalysts do not meet the requirements for a large-scale application and its properties have to be optimized. This includes not only the chemoselectivity, regioselectivity and stereoselectivity, but also long-term stability of the biocatalyst at certain temperatures or pH-values and activity in the presence of high substrate concentrations. Protein engineering has emerged as an important tool to overcome the limitations of natural enzymes as biocatalysts. There are two general strategies for protein engineering, rational design and directed evolution. In rational design detailed knowledge of the structure and function of the protein is used to make desired changes. Directed evolution involves either a random mutagenesis of the gene encoding the enzyme (e.g. by error-prone PCR) or recombination of gene fragments derived from DNase degradation, random priming recombination, random chimeragenesis on transient templates or recombined extension on truncated templates. In this review the essential methods for directed evolution of enzymes are described and various examples for the application of these protein engineering tools are provided.
1. Introduction. 2. Improvement of enzymes using non-recombinant methods. 3. Improvement of enzymes using gene-recombination methods. 4. Summary