- Kosol, Simone;
- Gallo, Angelo;
- Griffiths, Daniel;
- Valentic, Timothy R;
- Masschelein, Joleen;
- Jenner, Matthew;
- de los Santos, Emmanuel LC;
- Manzi, Lucio;
- Sydor, Paulina K;
- Rea, Dean;
- Zhou, Shanshan;
- Fülöp, Vilmos;
- Oldham, Neil J;
- Tsai, Shiou-Chuan;
- Challis, Gregory L;
- Lewandowski, Józef R
Modular polyketide synthases and non-ribosomal peptide synthetases are molecular assembly lines that consist of several multienzyme subunits that undergo dynamic self-assembly to form a functional megacomplex. N- and C-terminal docking domains are usually responsible for mediating the interactions between subunits. Here we show that communication between two non-ribosomal peptide synthetase subunits responsible for chain release from the enacyloxin polyketide synthase, which assembles an antibiotic with promising activity against Acinetobacter baumannii, is mediated by an intrinsically disordered short linear motif and a β-hairpin docking domain. The structures, interactions and dynamics of these subunits were characterized using several complementary biophysical techniques to provide extensive insights into binding and catalysis. Bioinformatics analyses reveal that short linear motif/β-hairpin docking domain pairs mediate subunit interactions in numerous non-ribosomal peptide and hybrid polyketide-non-ribosomal peptide synthetases, including those responsible for assembling several important drugs. Short linear motifs and β-hairpin docking domains from heterologous systems are shown to interact productively, highlighting the potential of such interfaces as tools for biosynthetic engineering.