Synthesis of¹³C/¹⁹F/²H labeled indoles for use as tryptophan precursors for protein NMR spectroscopy

Synthesis of indoles labeled with¹³C-¹H and¹³C-¹⁹F spin pairs is described. All syntheses utilize inexpensive carbon-¹³C dioxide as the¹³C isotope source. Ruthenium-mediated ring-closing metathesis is the key step in construction of the¹³C containing indole carbocycle. Fluorine is introducedviaelectrophilic fluorination at the 7-position andviapalladium-mediated cross-coupling at the 4-position. Indole and fluoroindoles are viable tryptophan precursors forin vivoprotein expression. We show that they are viable also inin vitroprotein synthesis using standardE. coliS30 extracts. Incorporation of the synthesized¹³C-¹H and¹³C-¹⁹F spin pair labeled tryptophans into proteins enables high-resolution and high-sensitivity nuclear magnetic resonance (NMR) spectroscopy.