Data-Driven Protein Engineering for Improving Catalytic Activity and Selectivity.

Protein engineering is essential for altering the substrate scope, catalytic activity and selectivity of enzymes for applications in biocatalysis. However, traditional approaches, such as directed evolution and rational design, encounter the challenge in dealing with the experimental screening process of a large protein mutation space. Machine learning methods allow the approximation of protein fitness landscapes and the identification of catalytic patterns using limited experimental data, thus providing a new avenue to guide protein engineering campaigns.

A Universal, Continuous Assay for SAM-dependent Methyltransferases.

Enzyme-catalyzed late-stage functionalization (LSF), such as methylation of drug molecules and lead structures, enables direct access to more potent active pharmaceutical ingredients (API). S-adenosyl-l-methionine-dependent methyltransferases (MTs) can play a key role in the development of new APIs, as they catalyze the chemo- and regioselective methylation of O-, N-, S- and C-atoms, being superior to traditional chemical routes. To identify suitable MTs, we developed a continuous fluorescence-based, high-throughput assay for SAM-dependent methyltransferases, which facilitates screening using E.

Structure- and Data-Driven Protein Engineering of Transaminases for Improving Activity and Stereoselectivity.

Amine transaminases (ATAs) are powerful biocatalysts for the stereoselective synthesis of chiral amines. Machine learning provides a promising approach for protein engineering, but activity prediction models for ATAs remain elusive due to the difficulty of obtaining high-quality training data. Thus, we first created variants of the ATA from Ruegeria sp.

Creation of ()-Amine Transaminase Activity within an α-Amino Acid Transaminase Scaffold.

The enzymatic transamination of ketones into ()-amines represents an important route for accessing a range of pharmaceuticals or building blocks. Although many publications have dealt with enzyme discovery, protein engineering, and the application of ()-selective amine transaminases [()-ATA] in biocatalysis, little is known about the actual role and how these enzymes have evolved from the ubiquitous α-amino acid transaminases (α-AATs). Here, we show the successful introduction of an ()-transaminase activity in an α-amino acid aminotransferase with one to six amino acid substitutions in the enzyme's active site.