A study of the reaction of glyceraldehyde with glyceraldehyde 3-phosphate dehydrogenase.

Kinetic data indicate that rabbit muscle glyceraldehyde d-phosphate dehydrogenase possesses two types of diphosphopyridine nucleotide-binding sites. The actual chemical reaction (oxidation-reduction) occurs at tight sites. Binding of DPN to loose sites activates the reaction at the tight sites. These results are consistent with the concept that the enzyme is “double headed,” with oxidation-reduction heads (which contain the tight sites) and transfer heads (which contain the loose sites). Light scattering data are consistent with the concept that (in the absence of arsenate or phosphate) the enzyme can exist as a dimer (with an apparent weight average molecular weight of 2.8 X 105) which can dissociate upon dilution into monomers (with molecular weights of 1.4 X 105). Dimerization is inhibited by high concentrations of arsenate (0.05 M). This inhibition is overcome by adding DPN to the loose sites. The monomer is unreactive in the absence of arsenate and is markedly activated by adding DPN to the loose sites. The dimer is reactive in the absence of arsenate and is only slightly activated by adding DPN to the loose sites. A kinetic mechanism is proposed which agrees with these and most other observations. This mechanism is sequential with the addition of DPN and then the aldehyde to the oxidation-reduction head. This is followed by the addition of arsenate and then DPN to the transfer head. The mech anism of the reaction with glyceraldehyde and glyceraldehyde 3-phosphate seems to be the same.