Abstract
A method for the purification of yeast K+-activated aldehyde dehydrogenase is presented which can be completed in substantially less time than other published procedures. The enzyme has a different N-terminal amino acid from preparations previously reported, and other small differences in amino acid content. These differences may be the result of differential proteolytic digestion rather than a different protein in vivo. A purification step involves the biospecific adsorption on affinity columns containing immobilized nucleotides in the absence of the substrate aldehyde. Direct binding studies with the coenzyme in the absence of aldehyde reveal 4 NAD sites per tetrameric molecule, each with a dissociation constant of 120 micron. These results conflict with properties of preparations previously reported and may conflict with kinetic models that have aldehyde as the leading substrate. Binding to Blue Dextran affinity columns suggests the presence of a dinucleotide fold in common with other dehydrogenases and kinases.
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