Reductases Produce Nitric Oxide in an Alternative Pathway to Form the Diazeniumdiolate Group of l-Alanosine

Abstract

l-Alanosine is a diazenium­diolate (N-nitrosohydroxylamine) antibiotic that inhibits MTAP-deficient tumor cells by blocking de novo adenine biosynthesis. Previous work revealed the early steps in the biosynthesis of l-alanosine. In the present study, we used genome mining to discover two new l-alanosine-producing strains that lack the aspartate-nitrosuccinate pathway genes found in the original l-alanosine producer. Instead, nitrate is reduced with a unique set of nitrate–nitrite reductases. These enzymes are typically used as part of the nitrogen cycle for denitrification or assimilation, and our report here shows how enzymes from the nitrogen cycle can be repurposed for the biosynthesis of specialized metabolites. The widespread distribution of nitric-oxide-producing reductases also indicates a potential for the discovery of new nitric-oxide-derived natural products.

Identification of the p-coumaric acid biosynthetic gene cluster in Kutzneria albida: insights into the diazotization-dependent deamination pathway

“…As a result, we discovered 134 BGCs and analyzed them using antiSMASH [ 17 ] and BiG-SCAPE (Figures S1 and S2 in Supporting Information File 1 ) [ 18 ]. Interestingly, approximately half of them (72 BGCs) do not have avaE and avaD homologs for the ANS pathway, suggesting three possibilities: (i) avaE and avaD homologs are encoded apart from the cluster, (ii) the strains have an alternative route to synthesize nitrous acid (as reported for alanosine biosynthesis [ 19 ]), and (iii) the strains use exogenous nitrous acid [ 13 ]. The BiG-SCAPE analysis divided the BGCs into seven groups (Figures S1 and S2 in Supporting Information File 1 ) and the ava cluster belongs to FAM_00127 (Figure S1).…”

Chalkophomycin Biosynthesis Revealing Unique Enzyme Architecture for a Hybrid Nonribosomal Peptide Synthetase and Polyketide Synthase

“…diazo, hydrozones, pyrazole, and diazeniumdiolate [43]. Pioneering studies of diazeniumdiolate biosynthesis in streptozotocin, L-alanosine, and fragin/valdiazen revealed multiple unique enzymes en route for N-N construction and further morphing from amino acid precursors, including SnzE/SznF, AlnDEFGLMN, HamACED/HamACEDG [3,[7][8][9][44][45]. In particular, Hertweck and co-workers recently discovered GrbED and their homologs responsible for biosynthesis of L-graminine, an unnatural amino acid found in a handful of cyclic or linear peptides, such as gramibactin, gladiobactin, JBIR-141/JBIR-142,megapolibactins, plantaribactin, and trinickiabactins [10][11].…”

Novel Biocatalysts from Specialized Metabolism

“…[38] Homologues of the enzymes CreD and CreE, which release nitrite after oxidation of Asp are also found in the biosynthesis of the amino acids alanosine (17) and graminin, which carry diazenium diolate side chains. [39][40][41] Streptozotocin ( 18) is an N-nitroso compound that is interesting due to its role in cancer chemotherapy. The loss of nitric oxide from the N-nitroso group contributes to the drug's bioactivity.…”