Structural investigation of the docking domain assembly from trans-AT polyketide synthases

Docking domains (DDs) located at the C- and N-termini of polypeptides play a crucial role in directing the assembly of polyketide synthases (PKSs), which are multienzyme complexes. Here, we determined the crystal structure of a complex comprising the C-terminal DD (CDDMlnB) and N-terminal DD (NDDMlnC) of macrolactin trans-acyltransferase (AT) PKS that were fused to a functional enzyme, AmpC EC2 β-lactamase. Interface analyses of the CDDMlnB/NDDMlnC complex revealed the molecular intricacies in the core section underpinning the precise DD assembly. Additionally, circular dichroism and steady-state kinetics demonstrated that the formation of the CDDMlnB/NDDMlnC complex had no influence on the structural and functional fidelity of the fusion partner, AmpC EC2. This inspired us to apply the CDDMlnB/NDDMlnC assembly to metabolon engineering. Indeed, DD assembly induced the formation of a complex between 4-coumarate-CoA ligase and chalcone synthase both involved in flavonoid biosynthesis, leading to a remarkable increase in naringenin production in vitro. [Display omitted] •Crystal structure of a docking domain (DD) complex fused to a functional enzyme•Structural determinants for the specificity between the cognate DD pairs revealed•Kd value for DDs fused to two distinct metabolic enzymes was determined•DD assembly yielded an approx. 2-fold increased production of naringenin Son et al. determined the crystal structure of the docking domains (DDs) complex from macrolactin trans-AT PKS, fused to a functional enzyme. The authors measured the Kd value for DDs fused to two distinct metabolic enzymes. DD-mediated substrate channeling was demonstrated by an increased naringenin production in flavonoid biosynthesis.