scholarly journals Redirecting RiPP biosynthetic enzymes to proteins and backbone-modified substrates

2022 ◽  
Author(s):  
Joshua A Walker ◽  
Noah Hamlish ◽  
Avery Tytla ◽  
Daniel D Brauer ◽  
Matthew B Francis ◽  
...  

Ribosomally synthesized and post-translationally modified peptides (RiPPs) are peptide-derived natural products that include the FDA-approved analgesic ziconotide1,2 as well as compounds with potent antibiotic, antiviral, and anticancer properties.3 RiPP enzymes known as cyclodehydratases and dehydrogenases represent an exceptionally well-studied enzyme class.3 These enzymes work together to catalyze intramolecular, interresidue condensation3,4 and aromatization reactions that install oxazoline/oxazole and thiazoline/thiazole heterocycles within ribosomally produced polypeptide chains. Here we show that the previously reported enzymes MicD-F and ArtGox accept backbone-modified monomers, including aramids and beta-amino acids, within leader-free polypeptides, even at positions immediately preceding or following the site of cyclization/dehydrogenation. The products are sequence-defined chemical polymers with multiple, diverse, non-alpha-amino acid subunits. We show further that MicD-F and ArtGox can install heterocyclic backbones within protein loops and linkers without disrupting the native tertiary fold. Calculations reveal the extent to which these heterocycles restrict conformational space; they also eliminate a peptide bond. Both features could improve the stability or add function to linker sequences now commonplace in emerging biotherapeutics. Moreover, as thiazoles and thiazoline heterocycles are replete in natural products,5,6,7 small molecule drugs,8,9 and peptide-mimetic therapeutics,10 their installation in protein-based biotherapeutics could improve or augment performance, activity, stability, and/or selectivity. This work represents a general strategy to expand the chemical diversity of the proteome beyond and in synergy with what can now be accomplished by expanding the genetic code.

2015 ◽  
Vol 81 (13) ◽  
pp. 4339-4350 ◽  
Author(s):  
Qi Zhang ◽  
James R. Doroghazi ◽  
Xiling Zhao ◽  
Mark C. Walker ◽  
Wilfred A. van der Donk

ABSTRACTLanthionine-containing peptides (lanthipeptides) are a rapidly growing family of polycyclic peptide natural products belonging to the large class of ribosomally synthesized and posttranslationally modified peptides (RiPPs). Lanthipeptides are widely distributed in taxonomically distant species, and their currently known biosynthetic systems and biological activities are diverse. Building on the recent natural product gene cluster family (GCF) project, we report here large-scale analysis of lanthipeptide-like biosynthetic gene clusters fromActinobacteria. Our analysis suggests that lanthipeptide biosynthetic pathways, and by extrapolation the natural products themselves, are much more diverse than currently appreciated and contain many different posttranslational modifications. Furthermore, lanthionine synthetases are much more diverse in sequence and domain topology than currently characterized systems, and they are used by the biosynthetic machineries for natural products other than lanthipeptides. The gene cluster families described here significantly expand the chemical diversity and biosynthetic repertoire of lanthionine-related natural products. Biosynthesis of these novel natural products likely involves unusual and unprecedented biochemistries, as illustrated by several examples discussed in this study. In addition, class IV lanthipeptide gene clusters are shown not to be silent, setting the stage to investigate their biological activities.


PLoS Biology ◽  
2020 ◽  
Vol 18 (12) ◽  
pp. e3001026
Author(s):  
Alexander M. Kloosterman ◽  
Peter Cimermancic ◽  
Somayah S. Elsayed ◽  
Chao Du ◽  
Michalis Hadjithomas ◽  
...  

Microbial natural products constitute a wide variety of chemical compounds, many which can have antibiotic, antiviral, or anticancer properties that make them interesting for clinical purposes. Natural product classes include polyketides (PKs), nonribosomal peptides (NRPs), and ribosomally synthesized and post-translationally modified peptides (RiPPs). While variants of biosynthetic gene clusters (BGCs) for known classes of natural products are easy to identify in genome sequences, BGCs for new compound classes escape attention. In particular, evidence is accumulating that for RiPPs, subclasses known thus far may only represent the tip of an iceberg. Here, we present decRiPPter (Data-driven Exploratory Class-independent RiPP TrackER), a RiPP genome mining algorithm aimed at the discovery of novel RiPP classes. DecRiPPter combines a Support Vector Machine (SVM) that identifies candidate RiPP precursors with pan-genomic analyses to identify which of these are encoded within operon-like structures that are part of the accessory genome of a genus. Subsequently, it prioritizes such regions based on the presence of new enzymology and based on patterns of gene cluster and precursor peptide conservation across species. We then applied decRiPPter to mine 1,295 Streptomyces genomes, which led to the identification of 42 new candidate RiPP families that could not be found by existing programs. One of these was studied further and elucidated as a representative of a novel subfamily of lanthipeptides, which we designate class V. The 2D structure of the new RiPP, which we name pristinin A3 (1), was solved using nuclear magnetic resonance (NMR), tandem mass spectrometry (MS/MS) data, and chemical labeling. Two previously unidentified modifying enzymes are proposed to create the hallmark lanthionine bridges. Taken together, our work highlights how novel natural product families can be discovered by methods going beyond sequence similarity searches to integrate multiple pathway discovery criteria.


2020 ◽  
Author(s):  
Jingxia Lu ◽  
Yuan Wu ◽  
Jiao Li ◽  
Yuqing Li ◽  
Yingying Zhang ◽  
...  

ABSTRACTThioamitides are a group of ribosomally synthesized and post-translational modified peptides with potent antiproliferative and pro-apoptotic activities. Their biosynthesis remains largely unknown, especially for the characteristic C-terminal 2-aminovinyl-Cysteine (AviCys) motifs. Herein, we report the discovery that homologs of class III lanthipeptide synthetases (LanKCts)encoded outside putative thioamitide biosynthetic gene clusters (BGCs) fully dehydrate the precursor peptides. Remarkably, LanKCt enzymes bind tightly to cysteine decarboxylases encoded inside thioamitide BGCs, and the resulting complex complete the macrocyclization of AviCys rings. Furthermore, LanKCt enzymes are present in the genomes of many thioamitide-producing strains and are functional when in complex with cysteine decarboxylases to produce AviCys macrocycles. Thus, our study reveals the participation of lanthipeptide synthetases as a general strategy for dehydration and AviCys formation during thioamitides biosynthesis and thus paves the way for the bioengineering of this class of bioactive natural products.


1990 ◽  
Vol 218 ◽  
Author(s):  
Peng Wang ◽  
Tara G. Hill ◽  
Mark D. Bednarski ◽  
Matthew R. Callstrom

The discovery of catalysts that can selectively couple unprotected peptide fragments would revolutionize protein chemistry by allowing convergent polypeptide synthesis. Proteolytic enzymes have the capability to perform this chemistry because the protein can specifically recognize and bind to C-terminal and N-terminal peptide sequences, activate the C-terminal peptide sequence by forming an acyl-enzyme intermediate, and couple the two peptide fragments together. However, barriers that limit the use of proteases as catalysts for convergent peptide synthesis include (i) the stability of proteolytic enzymes in organic solvent systems; (ii) a simple and effective C-erminal and N-terminal protecting group strategy; and (iii) the isolation of the polypeptide product from the reaction mixture. In the previous paper we reported the stabilization of enzymes by the covalent attachment of proteins through their ο-lysine residues to a series of carbohydrate-based macromolecules. In this paper we report the use of carbohydrate protein conjugates of proteases [CPC(proteases)] as catalysts for peptide bond synthesis and a general strategy for convergent oligopeptide synthesis.


2020 ◽  
Author(s):  
Katsuya Maruyama ◽  
Takashi Ishiyama ◽  
Yohei Seki ◽  
Kounosuke Oisaki ◽  
Motomu Kanai

A novel Tyr-selective protein bioconjugation using the water-soluble persistent iminoxyl radical is described. The conjugation proceeded with high Tyr-selectivity and short reaction time under biocompatible conditions (room temperature in buffered media under air). The stability of the conjugates was tunable depending on the steric hindrance of iminoxyl. The presence of sodium ascorbate and/or light irradiation promoted traceless deconjugation, restoring the native Tyr structure. The method is applied to the synthesis of a protein-dye conjugate and further derivatization to azobenzene-modified peptides.


2021 ◽  
Author(s):  
Sofie Dekimpe ◽  
Joleen Masschelein

Condensation domains perform highly diverse functions during natural product biosynthesis and are capable of generating remarkable chemical diversity.


Molecules ◽  
2021 ◽  
Vol 26 (3) ◽  
pp. 719
Author(s):  
Meri Yulvianti ◽  
Christian Zidorn

Cyanogenic glycosides are an important and widespread class of plant natural products, which are however structurally less diverse than many other classes of natural products. So far, 112 naturally occurring cyanogenic glycosides have been described in the phytochemical literature. Currently, these unique compounds have been reported from more than 2500 plant species. Natural cyanogenic glycosides show variations regarding both the aglycone and the sugar part of the molecules. The predominant sugar moiety is glucose but many substitution patterns of this glucose moiety exist in nature. Regarding the aglycone moiety, four different basic classes can be distinguished, aliphatic, cyclic, aromatic, and heterocyclic aglycones. Our overview covers all cyanogenic glycosides isolated from plants and includes 33 compounds with a non-cyclic aglycone, 20 cyclopentane derivatives, 55 natural products with an aromatic aglycone, and four dihydropyridone derivatives. In the following sections, we will provide an overview about the chemical diversity known so far and mention the first source from which the respective compounds had been isolated. This review will serve as a first reference for researchers trying to find new cyanogenic glycosides and highlights some gaps in the knowledge about the exact structures of already described compounds.


1966 ◽  
Vol 29 (3) ◽  
pp. 395-403 ◽  
Author(s):  
Takeshi Utsunomiya ◽  
Jay S. Roth

The RNase activity and properties of ribosome and polysome preparations from normal rat liver and some hepatomas have been examined. Polysome and ribosome preparations from the Novikoff, McCoy MDAB, and Dunning hepatomas had considerably higher specific RNase activity than corresponding preparations from normal rat liver, Novikoff ascites, or Morris 5123 hepatomas. The optimum pH of the RNase was approximately 8.5 for all samples tested, and the samples showed no evidence of latent RNase activity when treated with 3 M sodium chloride, EDTA, urea, or p-chloromercuribenzenesulfonic acid. The RNase activity appeared to be associated principally with breakdown products and/or subunits smaller than 80S. In the presence of Mg++ ions, subunits could reaggregate to form monomer ribosomes indistinguishable from the natural products, but some of the reassociated ribosomes could contain RNase activity which had been bound to the smaller particles. Similar results were obtained with spermine. In the hepatomas, evidence was obtained for the preexistence of considerable amounts of the smaller, RNase-containing subunits in the cell. When a small amount of crystalline bovine pancreatic RNase was added to partly dissociated ribosomes, the RNase was found only in association with the smaller subunits, and little or no enzyme was taken up by ribosomes or polysomes. The results have led to the conclusion that RNase is not a normal constituent of the ribosome or polysome, but that RNase may become associated with these particulates if dissociation and reassociation take place. Some implications of these findings for the stability of messenger RNA and for the mechanism of its breakdown are discussed.


Antioxidants ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 512
Author(s):  
Laurène Minsat ◽  
Cédric Peyrot ◽  
Fanny Brunissen ◽  
Jean-Hugues Renault ◽  
Florent Allais

The current cosmetic and nutraceutical markets are characterized by a strong consumer demand for a return to natural products that are less harmful to both the consumers and the environment than current petrosourced products. Phloretin, a natural dihydrochalcone (DHC) found in apple, has been widely studied for many years and identified as a strong antioxidant and anti-tyrosinase ingredient for cosmetic formulations. Its low concentration in apples does not allow it to be obtained by direct extraction from biomass in large quantities to meet market volumes and prices. Moreover, its remarkable structure prevents its synthesis through a green process. To overcome these issues, the synthesis of phloretin analogs appears as an alternative to access valuable compounds that are potentially more active than phloretin itself. Under such considerations, 12 chalcones (CHs) and 12 dihydrochalcones (DHCs) were synthesized through a green Claisen–Schmidt condensation using bio-based reagents. In order to evaluate the potential of these molecules, radical scavenging DPPH and anti-tyrosinase tests have been conducted. Moreover, the UV filtering properties and the stability of these analogs towards UV-radiations have been evaluated. Some molecules showed competitive antioxidant and anti-tyrosinase activities regarding phloretin. Two compounds in particular showed EC50 lower than phloretin, one chalcone and one dihydrochalcone.


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