Investigating docking domains in biosynthetic systems Munro Passmore 1 , Lona M. Alkhalaf 1 , Matthew Jenner 1,2 , Gregory L. Challis 1,2,3,4 1 Department of Chemistry, University of Warwick, UK, 2 Warwick Integrative Synthetic Biology Centre, University of Warwick, UK, 3 Department of Biochemistry and Molecular Biology, Monash University, Australia, 4 ARC Centre of Excellence for Innovations in Peptide and Protein Science, Monash University, Australia. Natural products biosynthesised by enzymatic assembly lines possess remarkable structural diversity and an array of antibiotic, anti-fungal and anti-cancer properties. Polyketide synthases (PKS) and non-ribosomal peptide synthetases (NRPS) represent two classes of these modular machines. Many of these biosynthetic pathways utilise docking domains to maintain the biosynthetic fidelity of interactions between assembly-line subunits. The strong and predictable interactions between docking domain partners guide their appended subunits together, assisting productive interaction. These are particularly interesting for potential bioengineering of assembly lines to produce novel bioactive products. Docking domains are especially critical in hybrid PKS-NRPS pathways, where the interface spans two very distinct types of assembly line. In this work, we present our investigation of the Short Linear Motif – β-Hairpin Docking Domain pair in the biosynthetic pathway of four related anti-cancer HDAC inhibitor products. Upon establishing an assay to probe the interaction between the cognate partner domains, we investigated the potential for cross-talk between systems across the PKS-NRPS interface. We will use this information to guide future engineering studies to introduce alternative NRPSs into the system. References 1. J. Masschelein, P.K. Sydor, C. Hobson, R. Howe, C. Jones, D.M. Roberts, Z.L. Yap, J. Parkhill, E. Mahenthiralingam and G.L. Challis. A dual transacylation mechanism for polyketide synthase chain release in enacyloxin antibiotic biosynthesis. Nat. Chem. 2019 , 11 , 906-912. 2. S. Kosol, A. Gallo, D. Griffiths, T.R. Valentic, J. Masschelein, M. Jenner, E.L. de los Santos, L. Manzi, P.K. Sydor, D. Rea, S. Zhou, V. Fülöp, N.J. Oldham, S.-C. Tsai, G.L. Challis, and J.R. Lewandowski. Structural basis for chain release from the enacyloxin polyketide synthase. Nat. Chem. 2019 , 11 , 913-923. 3. Vander Molen, K.M.; McCulloch, W.; Pearce, C.J.; Oberlies, N.H. Romidepsin (Istodax, NSC 630176, FR901228, FK228, Depsipeptide): A Natural Product Recently Approved for Cutaneous T-Cell Lymphoma. J. Antibiot. (Tokyo). 2011 , 64 (8), 525–531.
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