The DeLey-Doudoroff pathway of galactose metabolism in Azotobacter vinelandii

Abstract

Azotobacter vinelandii cell extracts reduced NAD+ and oxidized D- galactose to galactonate that subsequently was converted to 2-keto-3-deoxy- galactonate. Further metabolism of 2-keto-3-deoxy-galactonate required the presence of ATP and resulted in the formation of pyruvate and glyceraldehyde 3-P. Radiorespirometry indicated a preferential release of CO2 at the first carbon position of the D-galactose molecule. This suggested that Azotobacter vinelandii metabolizes D-galactose via the DeLey-Doudoroff pathway. The first enzyme of this pathway, D-galactose dehydrogenase, was partially characterized. It has a molecular weight of about 74,000 Da and an isoelectric point of 6.15. The pH optimum of the galactose dehydrogenase was about 9. The apparent K(m)s for NAD+ and D-galactose were 0.125 and 0.56 mM, respectively. Besides D-galactose, the active fraction of this galactose dehydrogenase also oxidized L-arabinose effectively. The electron acceptor for D-galactose or L-arabinose oxidation, NAD+, could not be replaced by NADP+. These substrate specificities were different from those reported in Pseudomonas saccharophila, Pseudomonas fluorescens, and Rhizobium meliloti.