Crystal Structure of the Acyltransferase Domain of the Iterative Polyketide Synthase in Enediyne Biosynthesis

Abstract

Background: DynE8 is an iterative polyketide synthase (PKS) that assembles polyketide intermediates from acetate units derived from malonyl-CoA. Results: We report the first acyltransferase (AT DYN10 ) crystal structure for an iterative PKS. Conclusion: AT DYN10 protects the malonyl-enzyme, but not the acetyl-enzyme intermediate, from hydrolysis and facilitates the transfer of malonyl to the acyl carrier protein.Significance: This differs from the dual specificity exhibited by acyltransferases of mammalian FAS and other iterative PKSs.

Reprogramming Acyl Carrier Protein Interactions of an Acyl-CoA Promiscuous trans-Acyltransferase

“…One of the top models was chosen on the basis of the alanine scanning mutagenesis data described above (Figure 6A). Overall, the docking model resembles that of the iterative type I PKS DynE8 AT and ACP (Liew, et al, 2012), and also that of the AT3-ACP3 model from DEBS (Wong, et al, 2010). ACP5 Kir is predicted to sit in a large cleft formed by parts of the KS-AT linker domain, as well as the small and large subdomains.…”

“…This is in agreement with phylogenetic analyses, which showed that KirCII is more closely related to cis-AT’s than to malonate-specific trans-ATs (Musiol and Weber, 2012). In addition to the large and small AT subdomains typical of trans-AT’s, the modeled KirCII structure also includes a KS-AT linker domain that is usually associated with cis-acting AT domains from type I PKSs (Liew, et al, 2012; Tang, et al, 2006). This prediction is in agreement with the sequence length of KirCII (which is longer than that of DSZS and MCAT), homology between the N-terminal portion of KirCII and various cis-AT’s (Figure S3), in addition to secondary structure similarity between the predicted KirCII KS-AT linker and structure of a known linker domain (Figure S4).…”

“…For example, the carboxylate side chain of D60 of ACP5 Kir is predicted to make a salt bridge with the terminus of the R410 side chain of KirCII (Figure 6A). Interestingly, docking models of the DynE8 AT:ACP (Liew, et al, 2012), DEBS AT4:ACP4 (Tang, et al, 2006), and fungal non-reducing PksA:ACP complexes (Bruegger, et al, 2013), also implicate an Asp equivalent to D60 of ACP5 Kir at the interface (Table S1). Notably, D60, along with the nearby D64, appear to form a negatively charged group of residues in loop-I (L 1 ) of ACP5 Kir that interacts with three consecutive Arg residues from KirCII (R408–R410).…”

“…Notably, D60, along with the nearby D64, appear to form a negatively charged group of residues in loop-I (L 1 ) of ACP5 Kir that interacts with three consecutive Arg residues from KirCII (R408–R410). Positively charged residues of other ATs are often involved in ACP interactions with an Asp residue analogous to ACP5 Kir D64 (Keatinge-Clay, et al, 2003; Liew, et al, 2012; Tang, et al, 2007; Tang, et al, 2006). In addition, ACP5 Kir R51 and R70, both judged as important for interaction with KirCII by alanine scanning mutagenesis, are positioned to interact with D98 and E395 of KirCII, respectively.…”

Structure-based analysis of the molecular interactions between acyltransferase and acyl carrier protein in vicenistatin biosynthesis

“…For example, S. coelicolor MCAT was suggested to interact with the loop region between helix I and II of actinorhodin PKS ACP (19,29). Alternatively, the presence of an additional domain such as the KS-AT linker domain in modular PKS might be involved in the binding mode of ACP, as proposed in the interaction between AT DYN10 and ACP DYN involved in dynemicin biosynthesis (30). For complete understanding of ACP recognition by AT, it is necessary to obtain other AT-ACP complex structures.…”