Biosynthesis of Strained Amino Acids Through a PLP-Dependent Enzyme via Cryptic Halogenation DOI Creative Commons
Max B. Sosa,

Jacob T. Leeman,

Lorenzo J. Washington

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2023, Volume and Issue: unknown

Published: Dec. 14, 2023

Abstract Amino acids (AAs) are modular and modifiable building blocks which nature uses to synthesize both macromolecules, such as proteins, small molecule natural products, alkaloids non-ribosomal peptides (NRPs). While the 20 main proteinogenic AAs display relatively limited side-chain diversity, a wide range of non-canonical amino (ncAAs) exist that not used by ribosome for protein synthesis but contain broad array structural features functional groups found in AAs. In this communication, we report discovery biosynthetic pathway new ncAA, pazamine, contains cyclopropane ring formed two steps. first step, chlorine is added onto C 4 position lysine radical halogenase PazA. The then next step pyridoxal-5’-phosphate-dependent enzyme, PazB, via an S N 2-like attack eliminate chloride. Genetic studies native host, Pseudomonas azotoformans , show pazamine its succinylated derivative, pazamide, potentially inhibit ethylene biosynthesis growing plants based on alterations root phenotype Arabidopsis thaliana seedlings. We further PazB can be utilized make alternative cyclobutane-containing AA. These discoveries may lead advances biocatalytic production specialty chemicals agricultural biotechnology.

Language: Английский

Biosynthesis of Strained Amino Acids by a PLP‐Dependent Enzyme through Cryptic Halogenation DOI
Max B. Sosa,

Jacob T. Leeman,

Lorenzo J. Washington

et al.

Angewandte Chemie International Edition, Journal Year: 2024, Volume and Issue: 63(31)

Published: March 23, 2024

Abstract Amino acids (AAs) are modular building blocks which nature uses to synthesize both macromolecules, such as proteins, and small molecule natural products, alkaloids non‐ribosomal peptides. While the 20 main proteinogenic AAs display relatively limited side chain diversity, a wide range of non‐canonical amino (ncAAs) exist that not used by ribosome for protein synthesis, but contain broad array structural features functional groups. In this communication, we report discovery biosynthetic pathway new ncAA, pazamine, contains cyclopropane ring formed in two steps. first step, chlorine is added onto C 4 position lysine radical halogenase, PazA. The then next step pyridoxal‐5′‐phosphate‐dependent enzyme, PazB, via an S N 2‐like attack at eliminate chloride. Genetic studies native host, Pseudomonas azotoformans , show pazamine potentially inhibits ethylene biosynthesis growing plants based on alterations root phenotype Arabidopsis thaliana seedlings. We further PazB can be utilized make alternative cyclobutane‐containing AA. These discoveries may lead advances biocatalytic production specialty chemicals agricultural biotechnology.

Language: Английский

Citations

5
Discovery of a Bacterial Hydrazine Transferase That Constructs the N-Aminolactam Pharmacophore in Albofungin Biosynthesis DOI
Wei Li, Ziyang Cheng,

Zhijie Zhao

et al.

Journal of the American Chemical Society, Journal Year: 2024, Volume and Issue: 146(19), P. 13399 - 13405

Published: May 3, 2024

Structural motifs containing nitrogen-nitrogen (N-N) bonds are prevalent in a large number of clinical drugs and bioactive natural products. Hydrazine (N

Language: Английский

Citations

5
Revision of the Formycin A and Pyrazofurin Biosynthetic Pathways Reveals Specificity for d-Glutamic Acid and a Cryptic N-Acylation Step During Pyrazole Core Formation DOI
Ziyang Zheng, Daan Ren, Yeonjin Ko

et al.

Journal of the American Chemical Society, Journal Year: 2025, Volume and Issue: unknown

Published: March 19, 2025

Formycin A and pyrazofurin are two naturally occurring pyrazole-derived C-nucleosides with antibacterial antiviral activities. While earlier studies have established the chemistry of C-glycosidic bond formation as well subsequent steps in biosynthesis formycin pyrazofurin, how pyrazole ring itself is constructed remains elusive. N-N was previously reported to involve coupling N6-hydroxylated l-lysine l-glutamic acid catalyzed by hydrazine synthetase PyfG, herein PyfG its homologue ForJ shown instead recognize d-glutamate l-glutamate. The product ForJ/PyfG catalysis then releases α-hydrazino d-glutamic upon processing NAD-dependent oxidoreductase ForL. Furthermore, N-acylation an amino ATP-grasp ligase ForM/PyfJ indispensable for recognition FAD-dependent ForR/PyfK perform dehydrogenation Cα-N thereby form a hydrazone intermediate. This work not only demonstrates that correct substrate but also reveals cryptic step assembly core. These results thus provide significant insights into rings rarely seen natural products.

Language: Английский

Citations

0
Synthetic Biology in Natural Product Biosynthesis DOI
Kaushik Seshadri,

Abner N. D. Abad,

Kyle K. Nagasawa

et al.

Chemical Reviews, Journal Year: 2025, Volume and Issue: unknown

Published: March 21, 2025

Synthetic biology has played an important role in the renaissance of natural products research during post-genomics era. The development and integration new tools have transformed workflow product discovery engineering, generating multidisciplinary interest field. In this review, we summarize recent developments biosynthesis from three different aspects. First, advances bioinformatics, experimental, analytical to identify associated with predicted biosynthetic gene clusters (BGCs) will be covered. This followed by extensive review on heterologous expression bacterial, fungal plant organisms. native host-independent paradigm identification, pathway characterization, enzyme is where synthetic most prominent role. Lastly, strategies engineer pathways for structural diversification complexity generation discussed, including assembly-line megasynthase precursor-directed modification, combinatorial biosynthesis.

Language: Английский

Citations

0
In vitro characterization of nonribosomal peptide synthetase-dependent O-(2-hydrazineylideneacetyl)serine synthesis indicates a stepwise oxidation strategy to generate the α-diazo ester moiety of azaserine DOI Creative Commons
Yusuke Shikai, Seiji Kawai, Yohei Katsuyama

et al.

Chemical Science, Journal Year: 2023, Volume and Issue: 14(33), P. 8766 - 8776

Published: Jan. 1, 2023

Azaserine, a natural product containing diazo group, exhibits anticancer activity. In this study, we investigated the biosynthetic pathway to azaserine. The putative azaserine gene (azs) cluster, which contains 21 genes, including those responsible for hydrazinoacetic acid (HAA) synthesis, was discovered using bioinformatics analysis of Streptomyces fragilis genome. Azaserine produced by heterologous expression azs cluster in albus. vitro enzyme assays recombinant Azs proteins revealed as follows. AzsSPTF and carrier protein (CP) AzsQ are used synthesize 2-hydrazineylideneacetyl (HDA) moiety attached from HAA. AzsD transfers HDA C-terminal CP domain AzsN. heterocyclization (Cy) nonribosomal peptide synthetase AzsO synthesizes O-(2-hydrazineylideneacetyl)serine (HDA-Ser) its l-serine moiety-attached thioesterase AzsB hydrolyzes it yield HDA-Ser, appears be converted oxidation. Bioinformatics Cy showed that has conserved DxxxxD motif; however, two amino residues (Thr Asp) important substituted Asn. Site-directed mutagenesis Asp motif (D193 D198) Asn (N414 N447) indicated these four ester bond synthesis. These results azasrine is synthesized stepwise oxidation HAA provided another strategy biosynthesize group.

Language: Английский

Citations

10
Bacterial Hydrazine Biosynthetic Pathways Featuring Cupin/Methionyl tRNA Synthetase‐like Enzymes DOI
Kenichi Matsuda, Toshiyuki Wakimoto

ChemBioChem, Journal Year: 2024, Volume and Issue: 25(9)

Published: March 9, 2024

Nitrogen-Nitrogen (N-N) bond-containing functional groups in natural products and synthetic drugs play significant roles exerting biological activities. The mechanisms of N-N bond formation organic molecules have garnered increasing attention over the decades. Recent advances illuminated various enzymatic nonenzymatic strategies, our understanding construction is rapidly expanding. A group didomain proteins with zinc-binding cupin/methionyl-tRNA synthetase (MetRS)-like domains, also known as hydrazine synthetases, generates amino acid-based hydrazines, which serve key biosynthetic precursors diverse functionalities such hydrazone, diazo, triazene, pyrazole, pyridazinone groups. In this review, we summarize current knowledge on pathways employing unique bond-forming machinery.

Language: Английский

Citations

3
Formation of Carboxymethyl-Phosphotriester Adducts in DNA DOI

Garrit Clabaugh,

Yinsheng Wang

Chemical Research in Toxicology, Journal Year: 2025, Volume and Issue: unknown

Published: April 16, 2025

Humans are exposed to endogenous and exogenous sources of N-nitroso compounds (NOCs). Metabolic activation some NOCs can yield diazoacetate, which is known induce the formation carboxymethylated DNA adducts that implicated in human gastrointestinal tumors. Although nucleobase have been investigated, no studies assessed if carboxymethylation occurs on phosphate backbone DNA. In this study, we report synthesis a carboxymethyl phosphotriester (CM-PTE) phosphoramidite building block thymidine preparation oligodeoxyribonucleotides (ODNs) containing site-specifically inserted CM-PTE. By employing liquid-chromatography-tandem mass spectrometry (LC-MS/MS) analysis, also demonstrated CM-PTE calf thymus treated with where identified total 16 products across all possible combinations flanking nucleobases. Together, our findings laid foundation for exploring vivo biological consequences lesions.

Language: Английский

Citations

0
Conserved Enzymatic Cascade for Bacterial Azoxy Biosynthesis DOI

Jingkun Shi,

Xin Zang,

Zhijie Zhao

et al.

Journal of the American Chemical Society, Journal Year: 2023, Volume and Issue: 145(49), P. 27131 - 27139

Published: Nov. 29, 2023

Azoxy compounds exhibit a wide array of biological activities and possess distinctive chemical properties. Although there has been considerable interest in the biosynthetic mechanisms azoxy metabolites, enzymatic basis responsible for bond formation remained largely enigmatic. In this study, we unveil enzyme cascade that constructs valanimycin biosynthesis. Our research demonstrates pair metalloenzymes, comprising membrane-bound hydrazine synthase nonheme diiron synthase, collaborate to convert an unstable pathway intermediate product through hydrazine-azo-azoxy pathway. Additionally, by characterizing homologues from other metabolite pathways, propose two-enzyme could represent conserved strategy bacteria. These findings provide significant mechanistic insights into N–N should facilitate targeted isolation bioactive genome mining.

Language: Английский

Citations

9
Phylogeny‐guided Characterization of Bacterial Hydrazine Biosynthesis Mediated by Cupin/methionyl tRNA Synthetase‐like Enzymes DOI
Kenichi Matsuda, Yuto Nakahara, Atina Rizkiya Choirunnisa

et al.

ChemBioChem, Journal Year: 2024, Volume and Issue: 25(7)

Published: Feb. 26, 2024

Cupin/methionyl-tRNA synthetase (MetRS)-like didomain enzymes catalyze nitrogen-nitrogen (N-N) bond formation between Nω-hydroxylamines and amino acids to generate hydrazines, key biosynthetic intermediates of various natural products containing N-N bonds. While the combination these two building blocks leads creation diverse hydrazine products, full extent their structural diversity remains largely unknown. To explore this, we herein conducted phylogeny-guided genome-mining related pathways consisting enzymes: flavin-dependent Nω-hydroxylating monooxygenases (NMOs) that produce Nω-hydroxylamine precursors cupin/MetRS-like couple with via A phylogenetic analysis identified unexplored sequence spaces enzyme families. The biochemical characterization NMOs demonstrated capabilities Nω-hydroxylamines, including those previously not known as Furthermore, five new novel combinations blocks, one non-amino acid blocks: 1,3-diaminopropane putrescine. This study substantially expanded variety forming mediated by enzymes.

Language: Английский

Citations

2
Biosynthesis and recruitment of reactive amino acids in nonribosomal peptide assembly lines DOI
Friedrich Johannes Ehinger, Christian Hertweck

Current Opinion in Chemical Biology, Journal Year: 2024, Volume and Issue: 81, P. 102494 - 102494

Published: June 26, 2024

Language: Английский

Citations

2