Five‐Membered Cyclitol Phosphate Formation by a myo‐Inositol Phosphate Synthase Orthologue in the Biosynthesis of the Carbocyclic Nucleoside Antibiotic Aristeromycin

Abstract

Aristeromycin is a unique carbocyclic nucleoside antibiotic produced by Streptomyces citricolor. In order to elucidate its intriguing carbocyclic formation, we used a genome-mining approach to identify the responsible enzyme. In silico screening with known cyclitol synthases involved in primary metabolism, such as myo-inositol-1-phosphate synthase (MIPS) and dehydroqunate synthase (DHQS), identified a unique MIPS orthologue (Ari2) encoded in the genome of S. citricolor. Heterologous expression of the gene cluster containing ari2 with a cosmid vector in Streptomyces albus resulted in the production of aristeromycin, thus indicating that the cloned DNA region (37.5 kb) with 33 open reading frames contains its biosynthetic gene cluster. We verified that Ari2 catalyzes the formation of a novel five-membered cyclitol phosphate from d-fructose 6-phosphate (F6P) with NAD as a cofactor. This provides insight into cyclitol phosphate synthase as a member of the MIPS family of enzymes. A biosynthetic pathway to aristeromycin is proposed based on bioinformatics analysis of the gene cluster.

Coordinated Biosynthesis of the Purine Nucleoside Antibiotics Aristeromycin and Coformycin in Actinomycetes

“…We also deduced that MacQ is likely responsible for self-resistance by deamination of ARM and NEP-A to alleviate the toxicity to host cells, as proposed previously (19). MacQ was expressed and purified from Escherichia coli (Fig.…”

“…Although S. citricolor is shown to be a COF producer in the present study, the genes for the biosynthesis of this antibiotic were missing in the surrounding region of the ari gene cluster ( Fig. 2A) (19), suggesting that they are probably distributed over the S. citricolor genome. To locate the COF gene cluster, we sequenced the genome of S. citricolor and used the conserved enzymes, including MacM (SAICAR synthetase) and MacN (short-chain dehydrogenase), as probes, allowing the identification of a candidate gene cluster (com) ( Fig.…”

“…Previous labeling studies have established that ARM is derived from D-glucose (14), and more recently, it was shown to arise from D-fructose-6-phosphate (F6P) (19). Investigation of the mac gene cluster led to the identification of 20 genes (except for macXMNO) that are involved in ARM biosynthesis.…”

“…Subsequently, Hill et al identified two key intermediates of the NEP-A and ARM biosynthetic pathway and tentatively proposed a concise pathway (18). Although the ARM gene cluster from Streptomyces citricolor NBRC 13005 (S. citricolor herein) has been identified and a plausible biosynthetic pathway has been proposed (19), several aspects remained to be determined.…”

Stereochemistry in the Reaction of the myo-Inositol Phosphate Synthase Ortholog Ari2 during Aristeromycin Biosynthesis

“…Aristeromycin effects its biological activity against bacteria, viruses, and tumor cells by inhibiting S -adenosyl- l -homocysteine hydrolase. − The compound exhibits the unique structural feature of a five-membered cyclitol moiety, which is connected to an adenine ring to form an adenosine mimic . Genome sequence analysis of the producer strain S. citricolor NBRC 13005 revealed that the myo -inositol phosphate synthase (MIPS) ortholog-containing biosynthetic gene cluster is responsible for the biosynthesis of aristeromycin . Furthermore, the MIPS ortholog Ari2 has been demonstrated to catalyze the formation of five-membered carbocyclic phosphate (2-hydroxymethyl)­cyclopentane-2,3,4,5-tetraol-1-phosphate ( 2 ) from the substrate d -fructose 6-phosphate (F6P) (Figure A) .…”