CysE and CysK, the last two enzymes of the cysteine biosynthetic pathway, engage in a bienzyme complex, cysteine synthase (CS), with yet incompletely characterized three-dimensional structure and regulatory function. Being absent in mammals, the two enzymes and their complex are attractive targets for antibacterial drugs. We have used hydrogen/deuterium exchange mass spectrometry to unveil how complex formation affects the conformational dynamics of CysK and CysE. Our results support a model where CysE is present in solution as a dimer of trimers, and each trimer can bind one CysK homodimer. When CysK binds to one CysE monomer, intra-trimer allosteric communication ensures conformational and dynamic symmetry within the trimer. Furthermore, a longer-range allosteric signal propagates through CysE to induce stabilization of the interface between the two CysE trimers, preparing the second trimer for binding the second CysK with a non-random orientation. These results provide new molecular insights into the allosteric formation of the CS complex and could help guide antibacterial drug design.