Fig. 2. Automated flow synthesis delivers diverse single-domain protein chains in both enantiomeric forms.

Analytical characterization of reverse phase-purified l- and d-chains from AFPS. Green traces show characterization of l-proteins, and blue traces show characterization of d-proteins. For each protein target, the following data are shown: (1) analytical HPLC trace of purified material recorded at 214 nm (bottom overlayed chromatograms); (2) integrated mass-to-charge spectrum of the total ion current (TIC) post-injection on a Q-TOF LC-MS instrument (separate insets on the left); and (3) the deconvolution of the TIC traces of (2) shown as overlayed insets on the right. Observed masses (abbreviated as Obs) from the deconvolution are shown along with the predicted values (abbreviated as Calc). The asterisks (*) in the analytical HPLC chromatograms indicate the major product peak corresponding to the integrated mass-to-charge and deconvoluted mass spectra depicted in the insets. The HPLC chromatograms for the l- and d-variants of barnase, BCL11a, Max-Max and Myc-Max dimers were acquired on different days, therefore variations in retention time were observed as a result of column conditioning. Previously reported structures are depicted for each of the targets, sourced from prior X-ray crystal structures available in the Protein Data Bank (PDB), NMR structures, or Alphafold predictions^50,51 (MDM2: 3FDO, ERG: 1SXE, Barnase: 1A2P, IRAK2: 3MOP, CHIP: 4KBQ, NEMO: 3BRV, FKBP12: 2PPN, BCL11a: 6KI6, YAP1: Alphafold prediction of Uniprot ID P46937 region 63–276, NEMO_iZIP: 6MI3, Myc-Max: 1HLO, Max-Max: 1NKP).