A model for the regulation of D-3-phosphoglycerate dehydrogenase, a Vmax-type allosteric enzyme

Escherichia coli D-3-phosphoglycerate dehydrogenase (ePGDH) is a tetramer of identical subunits that is allosterically inhibited by L-serine, the end product of its metabolic pathway. Because serine binding affects the velocity of the reaction and not the binding of substrate or cofactor, the enzyme is classified as of the Vmax type. Inhibition by a variety of amino acids and analogues of L-serine indicate that all three functional groups of serine are required for optimal interaction. Removing or altering any one functional group results in an increase in inhibitory concentration from micromolar to millimolar, and removal or alteration of any two functional groups removes all inhibitory ability. Kinetic studies indicate at least two serine-binding sites, but the crystal structure solved in the presence of bound serine and direct serine-binding studies show that there are a total of four serine-binding sites on the enzyme. However, approximately 85% inhibition is attained when only two sites are occupied. The three-dimensional structure of ePGDH shows that the serine-binding sites reside at the interface between regulatory domains of adjacent subunits. Two serine molecules bind at each of the two regulatory domain interfaces in the enzyme. When all four of the serines are bound, 100% inhibition of activity is seen. However, because the domain contacts are symmetrical, the binding of only one serine at each interface is sufficient to produce approximately 85% inhibition. The tethering of the regulatory domains to each other through multiple hydrogen bonds from serine to each subunit apparently prevents the body of these domains from undergoing the reorientation that must accompany a catalytic cycle. It is suggested that part of the conformational change may involve a hinge formed in the vicinity of the union of two antiparallel beta-sheets in the regulatory domains. The tethering function of serine, in turn, appears to prevent the substrate-binding domain from closing the cleft between it and the nucleotide-binding domain, which may be necessary to form a productive hydrophobic environment for hydride transfer. Thus, the structure provides a plausible model that is consistent with the binding and inhibition data and that suggests that catalysis and inhibition in this rare Vmax-type allosteric enzyme is based on the movement of rigid domains about flexible hinges.