Substrate Scope Expansion of Tryptophan Synthase for the Chemo‐Enzymatic Synthesis of Non‐Canonical Tryptophans

Tryptophan derivatives are valuable non-canonical amino acids widely used as precursors for the synthesis of bioactive molecules, as biophysical probes or introduced into polypeptides to improve or create entirely new functions. The enzyme tryptophan synthase (TrpS) provides a sustainable biocatalytic route to access these compounds directly from substituted indoles and l-serine. In this work, the substrate scope of Salmonella enterica tryptophan synthase (SeTrpS) was probed using a diverse library of synthetic indole derivatives bearing aryl, N-methylamino, alkynyl, and halogen substituents. Notably, SeTrpS displayed moderate to high conversions for most of the indoles tested, particularly for those substituted at positions 6 and 7. Two representative preparative scale reactions demonstrated the scalability of the process, affording the corresponding l-7-ethynyltryptophan and l-5-phenyltryptophan analogues in 58% and 25% yield, respectively, and excellent purity. This study contributes to illustrate the versatility of SeTrpS as a biocatalyst for the efficient and sustainable synthesis of structurally diverse tryptophan analogues, expanding the known substrate tolerance of this enzyme.