Incorporation of carbon atoms 1 and 6 of glucose into protocatechuic acid by Neurospora.

The mechanism of biosynthesis of the simpler aromatic compounds has been significantly clarified recently by the use of mutant strains of Neurospora (1) and Escherichia coli (2) and by the use of isotopic carbon (3-6). The evidence indicates that aromatization takes place via dehydroquinic acid, dehydroshikimic acid, and shikimic acid. Although the ultimate source of the carbon atoms of the aromatic rings of p-aminobenzoic acid, phenylalanine, tyrosine, and anthranilic acid is glucose, the mechanism by which t,hese carbon atoms form the basic 7-carbon unit of shikimic acid has not. yet been clearly established. The accumulation of protocatechuie acid by the aromaticless mutant st,rain of Neurospora crassa (1) seemed to provide a promising approach to this problem, since isotopic evidence had indicated that protocatechuic acid is derived from dehydroshikimic acid. Protocatcchuic acid would therefore be expected to have the same derivation and carbon labeling as dehydroshikimic acid, shikimic acid, and t,he aromatic nuclei of the amino acids. This paper rep0rt.s experiment.s in which protocatechuic acid produced by the a.romaticless mutant st.rain of Neurospora from glucose-l-C” and glucose-6-C*4 was degraded enzymatically and chemically via /%ketoadipic acid. The results show that carbon atoms 1 and 6 of glucose are incorporated almost exclusively, and to the exclusion of other glucose carbon atoms, int.o positions 2 and 6 of protocatcchuic acid. Posit.ion 2 is more strongly labeled from carbon atom 1 of glucose and posit,ion 6 from carbon atom 6. The results also indicat,c that in the production of protocatechuic acid the carbonyl oxygen at position 5 in dehydroshikimic acid is lost, and the two hydroxyl groups are retained. In the oxidative conversion of protocatechuic acid to fl-ketoadipic acid by Neurospora, the carbonyl oxygen appears to be introduced in position 6.