Ordered Binding of Substrates to Yeast Aldehyde Dehydrogenase

Yeast aldehyde dehydrogenase catalyzes the pyridine nucleotide-linked oxidation of a broad range of aldehydes to the corresponding acids. The reaction is experimentally irreversible. Initial velocity studies with DPN and two different aldehyde substrates, and equilibrium dialysis experiments with DPN, indicate that the substrates bind in a compulsory order; aldehyde and enzyme interact to make a binary complex which, in turn, forms a ternary complex with DPN. The equilibrium dialysis experiments were complicated by the ubiquitous presence of contaminating aldehydes and were made possible by the inclusion of a DPN-regenerating system. DPNH behaves as a competitive inhibitor of DPN and combines only with the enzyme aldehyde complex. Since neither of the pyridine nucleotides forms a binary complex with enzyme, the data fit a model in which aldehyde binds first while acid dissociates last.