Loading of KS Domains Using Acyl-ACPs

The psymberin polyketide core is the putative biosynthetic product of two genes psyA and psyD [17]. The first three PKS modules are encoded by psyA, which initiates the biosynthesis via an N-terminal GCN5-related A-acetyltransferase (GNAT) domain. GNAT domains catalyse the decarboxylation of malonyl-CoA to acetyl-CoA and subsequent loading to the downstream ACP [19]. Therefore, the native substrate of PsyA KSl is predicted to be acetyl-ACP. 

The substrate selectivity of PsyA KSl has been previously examined using SNAC-thioesters, with acetyl-SNAC being shown as the preferred substrate, despite a rather promiscuous substrate profile (see Sect. 3.2.3). It was postulated that the use of the corresponding acyl-ACPs would represent a more realistic approximation of the native system. Moreover, de-acylation of the small ACP domain could be monitored by mass spectrometry. The assay consisted of incubation of PsyA 

---