Coenzyme binding in crystals of glyceraldehyde-3-phosphate dehydrogenase.

Apo-glyceraldehyde-3-phosphate dehydrogenase from Bacillus stearothermophilus and the partially saturated holo-enzyme can be crystallized isomorphously with the entire tetramer occupying the crystal asymmetric unit. For crystals that contain one molecule of NAD+ per tetramer the coenzyme is bound uniquely in one of the four available sites. The presence of NAD+ gives rise to nonequivalence in the binding of a heavy-atom compound to the subunits of the tetramer while for the apo-enzyme this binding is clearly symmetric. These results suggest that NAD binding gives rise to sequential ligand-induced structural changes of the tetramer, which may be responsible for the observed negative cooperativity in coenzyme binding.