Biosynthesis of 3-Amino-5-hydroxybenzoic Acid, the Precursor of mC₇N Units in Ansamycin Antibiotics

Abstract

The biosynthetic pathway of 3-amino-5-hydroxybenzoic acid (AHBA) formation was studied with cell-free extracts from the rifamycin B producer, Amycolatopsis mediterranei S699, and the ansatrienin A producer, Streptomyces collinus Tü1892. Phosphoenolpyruvate (PEP) plus erythrose 4-phosphate (E4P) gave AHBA in low but nevertheless significant (6%) yield. 3,4-Dideoxy-4-amino-d-arabino-heptulosonic acid 7-phosphate (aminoDAHP) was converted efficiently into AHBA (45%), as were 5-deoxy-5-amino-3-dehydroquinic acid (aminoDHQ, 41%) and 5-deoxy-5-amino-3-dehydroshikimic acid (aminoDHS, 95%). On the other hand, the normal shikimate pathway intermediate, 3-deoxy-d-arabino-heptulosonic acid 7-phosphate (DAHP) did not give rise to AHBA under these conditions. AminoDAHP (9%) was produced by incubation of [14C]PEP and E4P, but not of [14C]DAHP, with the cell-free extracts. The results demonstrate the operation of a new variant of the shikimate pathway in the formation of the mC7N units of ansamycin, and presumably also mitomycin, antibiotics which leads from PEP, E4P, and a nitrogen source directly to aminoDAHP and then via aminoDHQ and aminoDHS to AHBA.

The biosynthesis of 3-amino-5-hydroxybenzoic acid (AHBA), the precursor of mC7N units in ansamycin and mitomycin antibiotics: a review

“…In reactions paralleling the normal shikimate pathway, aminoDAHP is then cyclized to 5-deoxy-5-aminodehydroquinic acid (aminoDHQ), dehydrated to 5-deoxy-5-aminodehydroshikimic acid (aminoDHS), and then aromatized to AHBA (Figure 2). The three postulated intermediates, all unknown compounds, were then synthesized 43,44 and incubated with cell-free extracts of the rifamycin producer, A. mediterranei. Their conversion into AHBA with high efficiency (aminoDAHP 45%, aminoDHQ 41%, aminoDHS B100%), but no conversion of DAHP, supported this hypothetical aminoshikimate pathway.…”

“…Their conversion into AHBA with high efficiency (aminoDAHP 45%, aminoDHQ 41%, aminoDHS B100%), but no conversion of DAHP, supported this hypothetical aminoshikimate pathway. 44 However, efforts to demonstrate the formation of aminoDAHP from E-4-P, PEP and glutamine in cell-free extracts of A. mediterranei or the ansatrienin-producer, Streptomyces collinus, gave only low yields of the product (up to 9%) and showed no dependence on the presence of glutamine. Furthermore, no significant incorporation of 15 N from [amide- 15 N]glutamine into AHBA was found.…”

“…These results, although supporting the later steps of the hypothetical pathway to AHBA, cast doubt on the postulated nature of the first step, the formation of aminoDAHP. 44 …”

A cybernetic model to predict the effect of freely available nitrogen substrate on rifamycin B production in complex media

“…The strain was cultivated and maintained as previously reported (Kim et al 1996). The seed culture was prepared in 100 ml medium containing glucose as a carbon source.…”

“…Fermentation was carried out in 250-ml flasks with single baffle (working volume 40 ml). Flasks were shaken at 240 RPM at 28°C (Kim et al 1996). All the fermentation experiments were performed in triplicate.…”