Höcker, B.; Lechner, H.

ChemBioChem 2021, 22, 1573-1577, 10.1002/cbic.202000880

An artificial cofactor based on an organocatalyst embedded in a protein has been used to conduct the Baylis-Hillman reaction in a buffered system. As protein host, we chose streptavidin, as it can be easily crystallized and thereby supports the design process. The protein host around the cofactor was rationally designed on the basis of high-resolution crystal structures obtained after each variation of the amino acid sequence. Additionally, DFT-calculated intermediates and transition states were used to rationalize the observed activity. Finally, repeated cycles of structure determination and redesign led to a system with an up to one order of magnitude increase in activity over the bare cofactor and to the most active proteinogenic catalyst for the Baylis-Hillman reaction known today.

Zeymer, C.

Protein Eng. Des. Sel. 2021, 34, 10.1093/protein/gzab003

Metalloproteins are essential to sustain life. Natural evolution optimized them for intricate structural, regulatory and catalytic functions that cannot be fulfilled by either a protein or a metal ion alone. In order to understand this synergy and the complex design principles behind the natural systems, simpler mimics were engineered from the bottom up by installing de novo metal sites in either natural or fully designed, artificial protein scaffolds. This review focuses on key challenges associated with this approach. We discuss how proteins can be equipped with binding sites that provide an optimal coordination environment for a metal cofactor of choice, which can be a single metal ion or a complex multinuclear cluster. Furthermore, we highlight recent studies in which artificial metalloproteins were engineered towards new functions, including electron transfer and catalysis. In this context, the powerful combination of de novo protein design and directed evolution is emphasized for metalloenzyme development.