The purification and properties of δ-aminolaevulic acid dehydrase

Abstract

The purification of an enzyme catalyzing the condensation of 6 -aminolaevulic acid to porphobilinogen is described. The enzyme, purified 270-fold when acetone-dried ox liver was used, had 1 component on electrophoresis at pH 8.4. The product of the reaction was isolated and identified as porphobilinogen, and evidence indicated no other Ehrlich-positive compound. The enzyme is widely distributed in nature and is probably present in all cells with an aerobic metabolism. In mammals, activity is highest in the liver, but kidney and bone marrow are also fairly active. The activity of liver appears to be confined to the soluble part of the cytoplasm. In anemia produced by phenylhydrazine the activity of the spleen and blood was increased 3-fold. Experimental porphyria produced by Sedormid was associated with a 2-fold rise in activity in the liver and kidney. After the 4th stage in the purification,the enzyme required cysteine or gluta-thione for activity. The activity of crude preparations which do not have this requirement was abolished by thiol inhibitors. Data strongly indicate that the enzyme requires thiol groups for activity. Ethylenedi-aminetetraacetate inhibited strongly, but no requirement for a metal was established. The enzyme was found to be inactive in the presence of aminotrishydroxymethylmethane under certain conditions; it did not act on aminoacetone, 6-amino-5-oxohexanoic acid and [alpha] [delta]-diamino-laevulic acid. The kinetics of the enzyme-catalysed reaction were in-vestigated. The pH optimum at 38[degree] is 6.7 and Km at pH 6.7 and 38[degree] is 1.4 x 10-4[image].