Extended Data Fig. 6: In vitro substrate specificity assay of AkaPKS4 and PheNRPS.
From: Plug-and-play engineering of modular polyketide synthases
a. Reaction schemes of AkaPKS4 with synthetic N-acetylcysteamine thioester (SNAC) substrates and the product structures of compounds 8b-9b. b. LC-MS analysis of AkaPKS4 with each substrate. 50 μL reaction mixture containing 10 mM ATP, 5 mM TCEP, 10 mM MgCl2, 1 mM CoA, 1 mM SNAC1-SNAC4, 1.6 mM malonate, 1 μM MatB, 400 mM PBS (pH 7.5), 2 μM AkaPKS4 were incubated at 30 °C overnight. Extracted ion chromatograms of m/z corresponding to the products were shown in purple (8b), orange (9a), blue (9b), respectively. c. Reaction schemes of PheNRPS with SNAC substrates and the product structures of compounds 11a–11d. d. LC-MS analysis of PheNRPS with each substrate. 50 μL reaction mixture containing 10 mM ATP, 5 mM TCEP, 10 mM MgCl2, 1 mM SNAC1-SNAC4, 1 mM alanine, 1 mM NADPH, 400 mM PBS (pH 7.5), 2 μM PheNRPS were incubated at 30 °C overnight. Extracted ion chromatograms of m/z corresponding to the products were shown in beige (11a), purple (11b), gray (11c), and blue (11d), respectively.
