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Coumermycin A1 is an aminocoumarin antibiotic produced by Streptomyces rishiriensis. It exhibits potent antibacterial and anticancer activity. The coumermycin A1 molecule contains two terminal 5-methyl-pyrrole-2-carboxylic acid moieties and one central 3-methylpyrrole-2,4-dicarboxylic acid moiety (CPM). While the biosynthesis of the terminal moieties has been elucidated in detail, the pathway leading to the CPM remains poorly understood. In this work, the minimal set of genes required for the generation of the CPM scaffold was identified. It comprises the five genes couR1, couR2a, couR2b, couR3, and couR4 which are grouped together in a contiguous 4.7 kb region within the coumermycin A1 biosynthetic gene cluster. The DNA fragment containing these genes was cloned into an expression plasmid and heterologously expressed in Streptomyces coelicolor M1146. Thereupon, the formation of CPM could be shown by HPLC and by HPLC-MS/MS, in comparison to an authentic CPM standard. This proves that the genes couR1–couR4 are sufficient to direct the biosynthesis of CPM, and that the adjacent genes couR5 and couR6 are not required for this pathway. The enzyme CouR3 was expressed in Escherichia coli and purified to near homogeneity. The protein exhibited an ATPase activity similar to that reported for its close ortholog, the threonine kinase PduX. However, we could not show a threonine kinase activity of CouR3, and; therefore, the substrate of CouR3 in CPM biosynthesis is still unknown and may be different from threonine.

The Chinese medicinal plant Artemisia annua L. (Annual Wormwood) contains the antimalarial compound artemisinin. The locally grown herb may offer an additional tool for the control of malaria, especially in poor countries where modern antimalarial drugs are often unavailable. In an open, randomized, controlled pilot trial, we investigated the efficacy and safety of traditional tea preparations of Artemisia annua in the treatment of uncomplicated malaria. Treatment resulted in a quick resolution of parasitaemia and of clinical symptoms. After 7 d of medication, cure rates were on average 74% for the Artemisia preparations compared with 91% for quinine. However, recrudescence rates were high in the Artemisia groups. Therefore, monotherapy with Artemisia annua L. cannot be recommended as alternative to modern antimalarials, but may deserve further investigation.

The bacterial gene ubiC encodes chorismate pyruvate-lyase, an enzyme which converts chorismate to 4-hydroxybenzoate and which is not normally present in plants. The UbiC protein was expressed in tobacco, with targeting of the gene product either to the plastids or to the cytosol. In both cases, chorismate pyruvate-lyase activity and a resulting formation of 4-hydroxybenzoate was detected. This suggests that chorismate, a metabolite of the shikimate pathway, is present not only in the plastids but also in the cytosol of plant cells.

A dimethylallyl diphosphate:4-hydroxyphenylpyruvate dimethylallyl transferase from the novobiocin producer Streptomyces spheroides DSM 40292 was identified, partially purified and characterized. The enzyme was soluble, and specific for 4-hydroxyphenylpyruvate and dimethylallyl diphosphate as substrates. It is most likely involved in the biosynthesis of the prenylated 4-hydroxybenzoate moiety of the aminocoumarin antibiotic novobiocin.

The ubiC gene of Escherichia coli encodes chorismate pyruvatelyase, an enzyme that converts chorismate into 4-hydroxybenzoate (4HB) and is not normally present in plants. The ubiC gene was expressed in Nicotiana tabacum L. plants under control of a constitutive plant promoter. The gene product was targeted into the plastid by fusing it to the sequence for the chloroplast transit peptide of the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase. Transgenic plants showed high chorismate pyruvate-lyase activity and accumulated 4HB as β-glucosides, with the glucose attached to either the hydroxy or the carboxyl function of 4HB. The total content of 4HB glucosides was approximately 0.52% of dry weight, which exceeded the content of untransformed plants by at least a factor of 1000. Feeding experiments with [1,7-13C2]shikimic acid unequivocally proved that the 4HB that was formed in the transgenic plants was not derived from the conventional phenylpropanoid pathway but from the newly introduced chorismate pyruvate-lyase reaction.

The enzymatic formation of p-hydroxybenzoate from p-coumarate in cell-free extracts of cell cultures of Lithospermum erythrorhizon Sieb. et Zucc. was investigated. p-Coumaroyl-coenzyme A (p-coumaroyl-CoA) is the activated intermediate in this biosynthetic reaction. It is formed by an ATP-, Mg2+ -, and CoA-dependent 4-hydroxycinnamate:CoA ligase reaction. p-Coumaroyl-CoA is oxidized and cleaved to p-hydroxybenzoyl-CoA and acetyl-CoA in a thioclastic reaction in which NAD is an essential cofactor. These CoA esters are rapidly hydrolyzed to acetate and p-hydroxybenzoate, probably by thioesterases. The enzymes involved in the formation of p-hydroxybenzoate are soluble. p-Hydroxybenzalde-hyde is not an intermediate in this conversion, and S-denosylmethionine and uridine-5[prime]-diphosphoglucose do not enhance formation of p-hydroxybenzoate in our system.