Allosteric activation of exopolysaccharide synthesis through cyclic di‐GMP‐stimulated protein–protein interaction

In many bacterial pathogens, the second messenger c‐di‐GMP stimulates the production of an exopolysaccharide (EPS) matrix to shield bacteria from assaults of the immune system. How c‐di‐GMP induces EPS biogenesis is largely unknown. Here, we show that c‐di‐GMP allosterically activates the synthesis of poly‐β‐1,6‐ N ‐acetylglucosamine (poly‐GlcNAc), a major extracellular matrix component of Escherichia coli biofilms. C‐di‐GMP binds directly to both PgaC and PgaD, the two inner membrane components of the poly‐GlcNAc synthesis machinery to stimulate their glycosyltransferase activity. We demonstrate that the PgaCD machinery is a novel type c‐di‐GMP receptor, where ligand binding to two proteins stabilizes their interaction and promotes enzyme activity. This is the first example of a c‐di‐GMP‐mediated process that relies on protein–protein interaction. At low c‐di‐GMP concentrations, PgaD fails to interact with PgaC and is rapidly degraded. Thus, when cells experience a c‐di‐GMP trough, PgaD turnover facilitates the irreversible inactivation of the Pga machinery, thereby temporarily uncoupling it from c‐di‐GMP signalling. These data uncover a mechanism of c‐di‐GMP‐mediated EPS control and provide a frame for c‐di‐GMP signalling specificity in pathogenic bacteria.