Extended Data Figure 5 : Partial mass spectra of active-site PikAIII and ACP4–PikAIII(C209A/ΔACP5) peptides from LC/FT-ICR MS of trypsin-digested proteins.

From: Structure of a modular polyketide synthase

Extended Data Figure 5

ad, ACP5 active-site peptides in their apo (a, b) and holo (with phosphopantetheine (Ppant) (c, d) states at 2+ and 3+ charge states. On the basis of integrated peak abundances from multiple LC/MS runs, greater than 97% of the ACP5 Ser 1438-containing peptides were modified with Ppant. e, f, Confirmation of the C209A mutation of the KS5 active site. The mutated active-site peptide was detected in the 4+ (e) and 3+ (f) charge states. gi, Example mass spectra of Ser 3605-containing active-site ACP4-derived peptides following enzymatic loading of the pentaketide from pentaketide–CoA. Both apo- (g), holo- (with Ppant) (h) and pentaketide–ACP4 (i) were detected. jl, Example mass spectra of active-site ACP4-derived peptides from a control experiment in which pentaketide–CoA was absent. The majority of the ACP4 active-site peptides were detected in the apo and holo states, while a very small percentage (<1%) contained the pentaketide intermediate. mp, ACP5 active-site peptides following incubation with MM–CoA. The MM building block was detected in high abundance on ACP5 Ser 1438 (o, p) with some unloaded holo-protein as well (m, n). qs, AT5 active-site peptides following incubation with MM–CoA. The MM building block was detected on AT Ser 655.