The Streptomyces master regulator BldD binds c-di-GMP sequentially to create a functional BldD2-(c-di-GMP)4 complex.

Streptomyces are ubiquitous soil bacteria that undergo a complex developmental transition coinciding with their production of antibiotics. This transition is controlled by binding of a novel tetrameric form of the second messenger， 3΄-5΄ cyclic diguanylic acid (c-di-GMP) to the master repressor， BldD. In all domains of life， nucleotide-based second messengers allow a rapid integration of external and internal signals into regulatory pathways that control cellular responses to changing conditions. c-di-GMP can assume alternative oligomeric states to effect different functions， binding to effector proteins as monomers， intercalated dimers or， uniquely in the case of BldD， as a tetramer. However， at physiological concentrations c-di-GMP is a monomer and little is known about how higher oligomeric complexes assemble on effector proteins and if intermediates in assembly pathways have regulatory significance. Here， we show that c-di-GMP binds BldD using an ordered， sequential mechanism and that BldD function necessitates the assembly of the BldD2-(c-di-GMP)4 complex.