scholarly journals Mutational Analysis of nocK and nocL in the Nocardicin A Producer Nocardia uniformis

2005 ◽  
Vol 187 (2) ◽  
pp. 739-746 ◽  
Author(s):  
Wendy L. Kelly ◽  
Craig A. Townsend
Keyword(s):  
Natural Products ◽  
Gene Cluster ◽  
Mutational Analysis ◽  
Biosynthetic Gene Cluster ◽  
Biosynthetic Gene ◽  
Wild Type ◽  
Nocardicin A ◽  
Lactam Ring ◽  
Undetermined Function

ABSTRACT The nocardicins are a family of monocyclic β-lactam antibiotics produced by the actinomycete Nocardia uniformis subsp. tsuyamanensis ATCC 21806. The most potent of this series is nocardicin A, containing a syn-configured oxime moiety, an uncommon feature in natural products. The nocardicin A biosynthetic gene cluster was recently identified and found to encode proteins in keeping with nocardicin A production, including the nocardicin N-oxygenase, NocL, in addition to genes of undetermined function, such as nocK, which bears similarities to a broad family of esterases. The latter was hypothesized to be involved in the formation of the critical β-lactam ring. While previously shown to effect oxidation of the 2′-amine of nocardicin C to provide nocardicin A, it was uncertain whether NocL was the only N-oxidizing enzyme required for nocardicin A biosynthesis. To further detail the role of NocL in nocardicin production in N. uniformis, and to examine the function of nocK, a method for the transformation of N. uniformis protoplasts to inactivate both nocK and nocL was developed and applied. A reliable protocol is reported to achieve both insertional disruption and in trans complementation in this strain. While the nocK mutant still produced nocardicin A at levels near that seen for wild-type N. uniformis, and therefore has no obvious role in nocardicin biosynthesis, the nocL disruptant failed to generate the oxime-containing metabolite. Nocardicin A production was restored in the nocL mutant upon in trans expression of the gene. Furthermore, the nocL mutant accumulated the biosynthetic intermediate nocardicin C, confirming its role as the sole oxime-forming enzyme required for production of nocardicin A.

scholarly journals mSphere of Influence: Synthetic Biology of Natural Product Biosynthesis

mSphere ◽  
2020 ◽  
Vol 5 (1) ◽  
Author(s):  
Mark C. Walker
Keyword(s):  
Natural Products ◽  
Synthetic Biology ◽  
Natural Product ◽  
Gene Cluster ◽  
Biosynthetic Gene Cluster ◽  
Biosynthetic Gene ◽  
Natural Product Biosynthesis ◽  
Antibiotic Compounds ◽  
Approaches To Study

ABSTRACT Mark Walker studies the biosynthesis and engineering of bacterial natural products with the long-term goal of identifying new antibiotic compounds. In this mSphere of Influence, he reflects on how “Direct cloning and refactoring of a silent lipopeptide biosynthetic gene cluster yields the antibiotic taromycin A” by K. Yamanaka, K. A. Reynolds, R. D. Kersten, K. S. Ryan, et al. (Proc Natl Acad Sci USA 111:1957–1962, 2014, https://doi.org/10.1073/pnas.1319584111) impacted his thinking on using synthetic biology approaches to study natural product biosynthesis.

scholarly journals Biosynthetic Potential of a Novel Antarctic Actinobacterium Marisediminicola antarctica ZS314T Revealed by Genomic Data Mining and Pigment Characterization

Marine Drugs ◽  
2019 ◽  
Vol 17 (7) ◽  
pp. 388 ◽  
Author(s):  
Li Liao ◽  
Shiyuan Su ◽  
Bin Zhao ◽  
Chengqi Fan ◽  
Jin Zhang ◽  
...  
Keyword(s):  
Data Mining ◽  
Natural Products ◽  
Gene Cluster ◽  
Genomic Data ◽  
Gene Clusters ◽  
Biosynthetic Gene Cluster ◽  
The Novel ◽  
Biosynthetic Gene ◽  
Base Pairs ◽  
Peptide Synthetase

Rare actinobacterial species are considered as potential resources of new natural products. Marisediminicola antarctica ZS314T is the only type strain of the novel actinobacterial genus Marisediminicola isolated from intertidal sediments in East Antarctica. The strain ZS314T was able to produce reddish orange pigments at low temperatures, showing characteristics of carotenoids. To understand the biosynthetic potential of this strain, the genome was completely sequenced for data mining. The complete genome had 3,352,609 base pairs (bp), much smaller than most genomes of actinomycetes. Five biosynthetic gene clusters (BGCs) were predicted in the genome, including a gene cluster responsible for the biosynthesis of C50 carotenoid, and four additional BGCs of unknown oligosaccharide, salinixanthin, alkylresorcinol derivatives, and NRPS (non-ribosomal peptide synthetase) or amino acid-derived compounds. Further experimental characterization indicated that the strain may produce C.p.450-like carotenoids, supporting the genomic data analysis. A new xanthorhodopsin gene was discovered along with the analysis of the salinixanthin biosynthetic gene cluster. Since little is known about this genus, this work improves our understanding of its biosynthetic potential and provides opportunities for further investigation of natural products and strategies for adaptation to the extreme Antarctic environment.

scholarly journals The Biosynthetic Gene Cluster for a Monocyclic β-Lactam Antibiotic, Nocardicin A

2004 ◽  
Vol 11 (7) ◽  
pp. 927-938 ◽  
Author(s):  
Michele Gunsior ◽  
Steven D Breazeale ◽  
Amanda J Lind ◽  
Jacques Ravel ◽  
James W Janc ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Biosynthetic Gene Cluster ◽  
Biosynthetic Gene ◽  
Nocardicin A ◽  
Lactam Antibiotic

scholarly journals Analysis of an Inactive Cyanobactin Biosynthetic Gene Cluster Leads to Discovery of New Natural Products from Strains of the Genus Microcystis

PLoS ONE ◽  
2012 ◽  
Vol 7 (8) ◽  
pp. e43002 ◽  
Author(s):  
Niina Leikoski ◽  
David P. Fewer ◽  
Jouni Jokela ◽  
Pirita Alakoski ◽  
Matti Wahlsten ◽  
...  
Keyword(s):  
Natural Products ◽  
Gene Cluster ◽  
Biosynthetic Gene Cluster ◽  
Biosynthetic Gene

scholarly journals Organization of the biosynthetic gene cluster for the macrolide concanamycin A in Streptomyces neyagawaensis ATCC 27449

Microbiology ◽  
2005 ◽  
Vol 151 (10) ◽  
pp. 3161-3169 ◽  
Author(s):  
Stephen F. Haydock ◽  
Anthony N. Appleyard ◽  
Tatiana Mironenko ◽  
John Lester ◽  
Natasha Scott ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Biosynthetic Pathway ◽  
Polyketide Synthase ◽  
Building Blocks ◽  
Biosynthetic Gene Cluster ◽  
Sequence Motif ◽  
Biosynthetic Gene ◽  
Concanamycin A ◽  
Modular Polyketide Synthase

The macrolide antibiotic concanamycin A has been identified as an exceptionally potent inhibitor of the vacuolar (V-type) ATPase. Such compounds have been mooted as the basis of a potential drug treatment for osteoporosis, since the V-ATPase is involved in the osteoclast-mediated bone resorption that underlies this common condition. To enable combinatorial engineering of altered concanamycins, the biosynthetic gene cluster governing the biosynthesis of concanamycin A has been cloned from Streptomyces neyagawaensis and shown to span a region of over 100 kbp of contiguous DNA. An efficient transformation system has been developed for S. neyagawaensis and used to demonstrate the role of the cloned locus in the formation of concanamycin A. Sequence analysis of the 28 ORFs in the region has revealed key features of the biosynthetic pathway, in particular the biosynthetic origin of portions of the backbone, which arise from the unusual polyketide building blocks ethylmalonyl-CoA and methoxymalonyl-ACP, and the origin of the pendant deoxysugar moiety 4′-O-carbamoyl-2′-deoxyrhamnose, as well as the presence of a modular polyketide synthase (PKS) encoded by six giant ORFs. Examination of the methoxymalonyl-specific acyltransferase (AT) domains has led to recognition of an amino acid sequence motif which can be used to distinguish methylmalonyl-CoA- from methoxymalonyl-ACP-specific AT domains in natural PKSs.

scholarly journals Discovery of the Pseudomonas Polyyne Protegencin by a Phylogeny-Guided Study of Polyyne Biosynthetic Gene Cluster Diversity

mBio ◽  
2021 ◽  
Author(s):  
Alex J. Mullins ◽  
Gordon Webster ◽  
Hak Joong Kim ◽  
Jinlian Zhao ◽  
Yoana D. Petrova ◽  
...  
Keyword(s):  
Biological Control ◽  
Natural Products ◽  
Gene Cluster ◽  
Plant Pathogens ◽  
Biological Activities ◽  
Pathogenic Fungi ◽  
Biosynthetic Gene Cluster ◽  
Biosynthetic Gene ◽  
Carbon Bond ◽  
Ecological Roles

Natural products bearing alkyne (triple carbon bond) or polyyne (multiple alternating single and triple carbon bonds) moieties exhibit a broad range of important biological activities. Polyyne metabolites have been implicated in important ecological roles such as cepacin mediating biological control of plant pathogens and caryoynencin protecting Lagriinae beetle eggs against pathogenic fungi.

scholarly journals Thioesterase from Cereulide Biosynthesis Is Responsible for Oligomerization and Macrocyclization of a Linear Tetradepsipeptide

2020 ◽  
Author(s):  
Christopher Boddy ◽  
Graham Heberlig
Keyword(s):  
Natural Products ◽  
Bacillus Cereus ◽  
Gene Cluster ◽  
Recent Work ◽  
Biosynthetic Gene Cluster ◽  
Alternative Mechanism ◽  
Biosynthetic Gene ◽  
Peptide Synthetases ◽  
Canonical Activity

Cereulide is a cyclic depsidodecapeptide produced in <i>Bacillus cereus</i> by two non-ribosomal peptide synthetases, CesA and CesB. While highly similar in structure and with a homologous biosynthetic gene cluster to valinomycin, recent work suggests that cereulide is produced via a different mechanism, which relys on a non-canonical coupling of two didepsipeptide-PCP bound intermediates. Ultimately this alternative mechanism generates a tetradepsipeptide-PCP bound intermediate that is prosed to differ from the tetradepsipeptide predicted from canonical activity of CesA and CesB. To test this hypothesis, we chemically synthetize both tetradepsipeptides as N-acetyl cysteamine thioesters and probed the ability of the purified recombinant terminal CesB thioesterase (CesB TE) to oligomerize and macrocyclize each substrate. Only the canonical substrate is converted cereulide, ruling out this alternative mechanism. We also show that CesB TE can use related tertradepsipeptide substrates, such as the valinomycin tetradespipetide and a hybride cereulide-valinomycin tetradespispetide in conjunction with its native substrate to generate chimeric natural products. This work clarifies the biosynthetic origins of cereulide and provides a powerful biocatalyst to access analogs of these ionophoric forming natural products.

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