scholarly journals Probing the promiscuity of ent-kaurene oxidases via combinatorial biosynthesis

2016 ◽  
Vol 113 (9) ◽  
pp. 2526-2531 ◽  
Author(s):  
Sibongile Mafu ◽  
Meirong Jia ◽  
Jiachen Zi ◽  
Dana Morrone ◽  
Yisheng Wu ◽  
...  
Keyword(s):  
Natural Products ◽  
Metabolic Engineering ◽  
Substrate Specificity ◽  
Plant Hormone ◽  
Structural Similarity ◽  
Combinatorial Biosynthesis ◽  
Underlying Structure ◽  
Engineering System ◽  
Insight Into

The substrate specificity of enzymes from natural products’ metabolism is a topic of considerable interest, with potential biotechnological use implicit in the discovery of promiscuous enzymes. However, such studies are often limited by the availability of substrates and authentic standards for identification of the resulting products. Here, a modular metabolic engineering system is used in a combinatorial biosynthetic approach toward alleviating this restriction. In particular, for studies of the multiply reactive cytochrome P450, ent-kaurene oxidase (KO), which is involved in production of the diterpenoid plant hormone gibberellin. Many, but not all, plants make a variety of related diterpenes, whose structural similarity to ent-kaurene makes them potential substrates for KO. Use of combinatorial biosynthesis enabled analysis of more than 20 such potential substrates, as well as structural characterization of 12 resulting unknown products, providing some insight into the underlying structure–function relationships. These results highlight the utility of this approach for investigating the substrate specificity of enzymes from complex natural products’ biosynthesis.

scholarly journals The crystal structure of mycobacterial epoxide hydrolase A

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Eike C. Schulz ◽  
Sara R. Henderson ◽  
Boris Illarionov ◽  
Thomas Crosskey ◽  
Stacey M. Southall ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Substrate Specificity ◽  
Drug Targets ◽  
Epoxide Hydrolase ◽  
Sequence Similarity ◽  
Structural Similarity ◽  
High Sequence Similarity ◽  
High Structural Similarity ◽  
Potential Drug Targets ◽  
Insight Into

Abstract The human pathogen Mycobacterium tuberculosis is the causative agent of tuberculosis resulting in over 1 million fatalities every year, despite decades of research into the development of new anti-TB compounds. Unlike most other organisms M. tuberculosis has six putative genes for epoxide hydrolases (EH) of the α/β-hydrolase family with little known about their individual substrates, suggesting functional significance for these genes to the organism. Due to their role in detoxification, M. tuberculosis EH’s have been identified as potential drug targets. Here, we demonstrate epoxide hydrolase activity of M. thermoresistibile epoxide hydrolase A (Mth-EphA) and report its crystal structure in complex with the inhibitor 1,3-diphenylurea at 2.0 Å resolution. Mth-EphA displays high sequence similarity to its orthologue from M. tuberculosis and generally high structural similarity to α/β-hydrolase EHs. The structure of the inhibitor bound complex reveals the geometry of the catalytic residues and the conformation of the inhibitor. Comparison to other EHs from mycobacteria allows insight into the active site plasticity with respect to substrate specificity. We speculate that mycobacterial EHs may have a narrow substrate specificity providing a potential explanation for the genetic repertoire of epoxide hydrolase genes in M. tuberculosis.

scholarly journals Functional Characterization of an Anthocyanin Dimalonyltransferase in Maize

Molecules ◽  
2021 ◽  
Vol 26 (7) ◽  
pp. 2020
Author(s):  
Michael Paulsmeyer ◽  
John Juvik
Keyword(s):  
Substrate Specificity ◽  
Structure Function ◽  
Functional Characterization ◽  
Malonyl Coa ◽  
Natural Colorants ◽  
Acylated Anthocyanins ◽  
The Stability ◽  
Kinetics Of ◽  
Insight Into

Anthocyanins are pigments with appealing hues that are currently being used as sources of natural colorants. The interaction of acylation on the stability of anthocyanin molecules has long been known. Maize is an abundant source of malonylglucoside and dimalonylglucoside anthocyanins. The enzyme Aat1 is an anthocyanin acyltransferase known to synthesize the majority of acylated anthocyanins in maize. In this paper, we characterize the substrate specificity and reaction kinetics of Aat1. It was found that Aat1 has anthocyanin 3-O-glucoside dimalonyltransferase activity and is only the second enzyme of this type characterized to this date. Our results indicate that Aat1 can utilize malonyl-CoA; succinyl-CoA and every anthocyanin 3-O-glucoside tested. Results of this study provide insight into the structure–function relations of dimalonyltransferases and give a unique insight into the activity of monocot anthocyanin acyltransferases.

scholarly journals A Fresh Insight into the Interaction of Natural Products with Pregnane X Receptor

2008 ◽  
Vol 3 (12) ◽  
pp. 1934578X0800301
Author(s):  
Salvador Máñez
Keyword(s):  
Natural Products ◽  
Biochemical Characterization ◽  
Pregnane X Receptor ◽  
Synthetic Drugs ◽  
Enzymatic Induction ◽  
Conjugating Enzymes ◽  
Fresh Insight ◽  
Insight Into ◽  
Endogenous Steroids

The discovery that various drugs (e.g., phenobarbital) stimulate their own metabolism through a mechanism coined as enzymatic induction opened up a fascinating road that eventually led to the accurate biochemical characterization of the pregnane X receptor. After numerous studies, researchers have concluded that this receptor is activated by different endogenous steroids and a number of foreign lipophile ligands. Once activated, it induces the synthesis of oxygenases and conjugating enzymes. The activating ligands identified to date include many synthetic drugs, along with a number of natural products. The present review summarizes the data relating to the origin, chemistry, and pharmacological activity of the newest natural products that have been found to interact with the pregnane X receptor.

scholarly journals Accessing Nature’s diversity through metabolic engineering and synthetic biology

F1000Research ◽  
2016 ◽  
Vol 5 ◽  
pp. 397 ◽  
Author(s):  
Jason R. King ◽  
Steven Edgar ◽  
Kangjian Qiao ◽  
Gregory Stephanopoulos
Keyword(s):  
Natural Products ◽  
Metabolic Engineering ◽  
Synthetic Biology ◽  
Natural Product ◽  
Chemical Space ◽  
Single Step ◽  
Enzyme Engineering ◽  
Biologically Active ◽  
Combinatorial Biosynthesis ◽  
Pathway Engineering

In this perspective, we highlight recent examples and trends in metabolic engineering and synthetic biology that demonstrate the synthetic potential of enzyme and pathway engineering for natural product discovery. In doing so, we introduce natural paradigms of secondary metabolism whereby simple carbon substrates are combined into complex molecules through “scaffold diversification”, and subsequent “derivatization” of these scaffolds is used to synthesize distinct complex natural products. We provide examples in which modern pathway engineering efforts including combinatorial biosynthesis and biological retrosynthesis can be coupled to directed enzyme evolution and rational enzyme engineering to allow access to the “privileged” chemical space of natural products in industry-proven microbes. Finally, we forecast the potential to produce natural product-like discovery platforms in biological systems that are amenable to single-step discovery, validation, and synthesis for streamlined discovery and production of biologically active agents.

Skeletal elements of crinoid echinoderms: High-resolution structural and microanalytical results

1983 ◽  
Vol 41 ◽  
pp. 194-195
Author(s):  
D. F. Blake ◽  
L. F. Allard ◽  
D. R. Peacor
Keyword(s):  
High Resolution ◽  
Marine Invertebrates ◽  
Sea Urchins ◽  
Structural Features ◽  
Sea Stars ◽  
High Resolution Tem ◽  
Relationship Of ◽  
The Relationship ◽  
Insight Into

Echinodermata is a phylum of marine invertebrates which has been extant since Cambrian time (c.a. 500 m.y. before the present). Modern examples of echinoderms include sea urchins, sea stars, and sea lilies (crinoids). The endoskeletons of echinoderms are composed of plates or ossicles (Fig. 1) which are with few exceptions, porous, single crystals of high-magnesian calcite. Despite their single crystal nature, fracture surfaces do not exhibit the near-perfect {10.4} cleavage characteristic of inorganic calcite. This paradoxical mix of biogenic and inorganic features has prompted much recent work on echinoderm skeletal crystallography. Furthermore, fossil echinoderm hard parts comprise a volumetrically significant portion of some marine limestones sequences. The ultrastructural and microchemical characterization of modern skeletal material should lend insight into: 1). The nature of the biogenic processes involved, for example, the relationship of Mg heterogeneity to morphological and structural features in modern echinoderm material, and 2). The nature of the diagenetic changes undergone by their ancient, fossilized counterparts. In this study, high resolution TEM (HRTEM), high voltage TEM (HVTEM), and STEM microanalysis are used to characterize tha ultrastructural and microchemical composition of skeletal elements of the modern crinoid Neocrinus blakei.

NMR characterization of complex natural products: Assigning novel, proton-deficient alkaloid scaffolds

Planta Medica ◽  
2016 ◽  
Vol 81 (S 01) ◽  
pp. S1-S381
Author(s):  
KR Gustafson ◽  
STS Chan ◽  
D Milanowski
Keyword(s):  
Natural Products ◽  
Nmr Characterization

Expanding the Natural Products Heterologous Expression Repertoire in the Model Cyanobacterium Anabaena sp. Strain PCC 7120: Production of Pendolmycin and Teleocidin B-4

2019 ◽  
Author(s):  
Patrick Videau ◽  
Kaitlyn Wells ◽  
Arun Singh ◽  
Jessie Eiting ◽  
Philip Proteau ◽  
...  
Keyword(s):  
Natural Products ◽  
Genome Mining ◽  
Gene Clusters ◽  
Combinatorial Biosynthesis ◽  
Test Case ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Cyanobacterium Anabaena ◽  
Anabaena Sp ◽  
Pcc 7120

Cyanobacteria are prolific producers of natural products and genome mining has shown that many orphan biosynthetic gene clusters can be found in sequenced cyanobacterial genomes. New tools and methodologies are required to investigate these biosynthetic gene clusters and here we present the use of <i>Anabaena </i>sp. strain PCC 7120 as a host for combinatorial biosynthesis of natural products using the indolactam natural products (lyngbyatoxin A, pendolmycin, and teleocidin B-4) as a test case. We were able to successfully produce all three compounds using codon optimized genes from Actinobacteria. We also introduce a new plasmid backbone based on the native <i>Anabaena</i>7120 plasmid pCC7120ζ and show that production of teleocidin B-4 can be accomplished using a two-plasmid system, which can be introduced by co-conjugation.

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