The metabolism of protocatechuic acid by Neurospora.

A mutant strain of Neurospora crassa, Y7655a, which requires for growth either phenylalanine, tyrosine, tryptophan, and p-aminobenzoic acid in combination or shikimic acid alone, was found to be blocked in the conversion of dehydroshikimic acid t.o shikimic acid. Some dehydroshikimic acid was accumulated during growth, but the major accumulated product was protocat.echuic acid (3,4-dihydroxybenzoic acid) (1). Characteristically, this strain quantitatively metabolizes its accumulated protocatechuic acid after exhaustion of the glucose or sucrose provided as the original carbon source. This phenomenon seemed of int.erest because of its apparent similarity to t.he “diauxie” frequently observed in inducible enzyme systems (2). Furthermore, it allowed an extension of the investigations of the metabolism of prot,ocatechuic acid heretofore carried out with bacteria (3-5) to Neurospora. This paper is primarily concerned with the biochemical transformations involved in the oxidation of protocatechuic acid to 8-ketoadipic acid. We have succeeded in separating the enzymes involved. The first, protocatechuic acid oxidase, which catalyzes the oxidation of protoeatechuic acid to /?-carboxymuconic acid (4), has been demonstrated to be present in extracm of the mutant strain. Little or no enzyme could be found in extracts of the wild type unless it had been exposed to protocatechuic acid or an inducing analogue during growth. The second, a lactonizing enzyme, converts ,8-carboxymuconic acid to its corresponding lactone which has been identified as (-)-&carboxy-y-carboxymethyl-A”but,enolide (B-carboxymuconolactone). The enzyme system responsible for the conversion of p-carboxymuconolactonc to fl-kctoadipic acid will be referred to as the