Insights into the function of trans-acyl transferase polyketide synthases from the SAXS structure of a complete module

Abstract

Combined analysis by SAXS, NMR and homology modeling reveals the structure of an apo module from a trans-acyltransferase polyketide synthase.

The structural biology of biosynthetic megaenzymes

“…We need, for example, corroborating structural data on PKS and NRPS modules of identical composition to those studied to date, ideally from several different systems 61,84,[95][96][97] ; these could, in the case of the cis-AT PKSs, allow for discrimination between the cryo-EM-and SAXS-derived models 95,97 . We also require information on intact modules of different domain composition to understand how such variations are accommodated.…”

“…57). Nonetheless, the structures of all three of the core domains have now been reported: the KS domain (one from the rhizoxin pathway solved as a didomain with a downstream branching (B) domain 58 and another monodomain KS fragment from the bacillaene PKS 59 ), a discrete AT domain 57 and multiple ACP domains 60,61 . In addition, structures of domains for several unusual tailoring activities from the bacillaene assembly line (a KR that reduces both α-and β-keto groups 62 and an enoyl-isomerase (EI) that catalyzes a double-bond shift 63 ) have been reported.…”

“…This observation has important implications for the length and flexibility of the linker required to join β-modification modules to downstream modules in engineered systems. Indeed, such linkers in trans-AT PKS systems are on average significantly longer than the corresponding linkers following nonmodifying modules (~63 versus 46 residues, respectively) 61 . The final result of this study was to reveal sequences that might function as C-and N-terminal docking domains.…”

“…SAXS was also used to structurally characterize an apo module from the virginiamycin trans-AT PKS 61 incorporates only essential domains (a KS and a tandem of ACPs characteristic of such β-methylation modules) and so was expected to have a structure distinct from modules that include tailoring activities. The SAXS analysis nonetheless revealed an overall architecture globally similar to that of Pik module 5, with the KS homodimer at the base of an arch and the two ACPs and their intervening linkers forming the walls (Fig.…”

The Combinatorial Biosynthesis of “Unnatural” Products with Polyketides

“…Technologies such as tandem equilibrium isotope exchange [68], electron cryo-microscopy [69], small-angle X-ray scattering (SAXS) [70,71], and bottom-up liquid chromatography with Fourier transform ion cyclotron resonance mass spectrometry [72] are of great help. More attention has been paid to the studies of PKS on a systematical level either in vivo or vitro, further accelerating the pace in the rational engineering of polyketide assembly lines [70,73]. In addition, the total enzymatic reconstruction of PKS biosynthesis pathway shows promising use in PKS biocatalytic analysis [74].…”