Cloning and Heterologous Expression of Three Type II PKS Gene Clusters fromStreptomyces bottropensis

ChemBioChem ◽  
2011 ◽  
Vol 13 (2) ◽  
pp. 224-230 ◽  
Author(s):  
Xiaohui Yan ◽  
Katharina Probst ◽  
Anton Linnenbrink ◽  
Moritz Arnold ◽  
Thomas Paululat ◽  
...  
Keyword(s):  
Heterologous Expression ◽  
Gene Clusters ◽  
Type Ii ◽  
Type Ii Pks

Identification of the kinanthraquinone biosynthetic gene cluster by expression of an atypical response regulator

2021 ◽  
Vol 85 (3) ◽  
pp. 714-721
Author(s):  
Risa Takao ◽  
Katsuyuki Sakai ◽  
Hiroyuki Koshino ◽  
Hiroyuki Osada ◽  
Shunji Takahashi
Keyword(s):  
Heterologous Expression ◽  
Gene Cluster ◽  
Secondary Metabolite ◽  
Response Regulator ◽  
Bac Library ◽  
Gene Clusters ◽  
Biosynthetic Gene Cluster ◽  
Gene Organization ◽  
Biosynthetic Gene ◽  
Streptomyces Sp

ABSTRACT Recent advances in genome sequencing have revealed a variety of secondary metabolite biosynthetic gene clusters in actinomycetes. Understanding the biosynthetic mechanism controlling secondary metabolite production is important for utilizing these gene clusters. In this study, we focused on the kinanthraquinone biosynthetic gene cluster, which has not been identified yet in Streptomyces sp. SN-593. Based on chemical structure, 5 type II polyketide synthase gene clusters were listed from the genome sequence of Streptomyces sp. SN-593. Among them, a candidate gene cluster was selected by comparing the gene organization with grincamycin, which is synthesized through an intermediate similar to kinanthraquinone. We initially utilized a BAC library for subcloning the kiq gene cluster, performed heterologous expression in Streptomyces lividans TK23, and identified the production of kinanthraquinone and kinanthraquinone B. We also found that heterologous expression of kiqA, which belongs to the DNA-binding response regulator OmpR family, dramatically enhanced the production of kinanthraquinones.

scholarly journals Discovery and Biosynthesis of Natural Products from New Zealand Soil Metagenome Libraries

2021 ◽  
Author(s):  
◽  
Luke Stevenson
Keyword(s):  
Natural Products ◽  
New Zealand ◽  
Heterologous Expression ◽  
Gene Clusters ◽  
Bioinformatic Analysis ◽  
Coli Strain ◽  
Functional Screening ◽  
Functional Difference ◽  
Biosynthetic Pathways ◽  
New Members

<p>Antibiotic discovery rates dramatically declined following the “golden age” of the 1940’s to the 1960’s. The platforms that underpinned that age of discovery rested upon laboratory cultivation of a small clade of bacteria, the actinomycetes, primarily isolated from soil environments. Fermentation extracts of these isolated bacteria have provided the majority of antibiotics and anticancer small molecules still used today. By applying modern genetic analysis techniques to these same environmental sources that have previously yielded such success, we can uncover new biosynthetic pathways, and bioactive compounds. The work described in this thesis investigated New Zealand soil metagenomes for this purpose.  Four large metagenome libraries were constructed from the microbiomes of diverse soil environments. These were then interrogated by a functional screening approach in a knockout Escherichia coli strain, to recover a large collection of the biosynthetic gene clusters responsible for bacterial secondary metabolite production. Using different modes of bioinformatic analysis, these gene clusters were demonstrated to have both phylogenetic divergence, and functional difference from bacterial biosynthesis pathways previously discovered from culture based studies.  Two additional biosynthetic pathways were recovered from one of these metagenome libraries, and in each case found to have novel genetic features. These gene clusters were further studied by heterologous expression within Streptomyces albus production hosts. One of these gene clusters produced small aromatic polyketide compounds, the structure of one of which was solved by chemical analytic techniques, and found to be a new chemical entity.  The second gene cluster was demonstrated to have similarity to known aureolic acid biosynthesis gene clusters – a class of potent anticancer natural products. Heterologous expression resulted in the production of many metabolites, two of which were characterised and found to be new members of this chemical class.  The research in this thesis both validates the use of metagenomic analysis for future natural product discovery efforts, and adds to a growing body of evidence that understudied clades of bacteria have an untapped biosynthetic potential that can be accessed by metagenomic methods.</p>

scholarly journals Enzyme-constrained models and omics analysis of Streptomyces coelicolor reveal metabolic changes that enhance heterologous production

2019 ◽  
Author(s):  
Snorre Sulheim ◽  
Tjaša Kumelj ◽  
Dino van Dissel ◽  
Ali Salehzadeh-Yazdi ◽  
Chao Du ◽  
...  
Keyword(s):  
Heterologous Expression ◽  
Streptomyces Coelicolor ◽  
Gene Clusters ◽  
Metabolic Model ◽  
Heterologous Production ◽  
Comprehensive Picture ◽  
Constrained Models ◽  
Systemic Understanding ◽  
Genome Scale ◽  
Further Development

AbstractMany biosynthetic gene clusters (BGCs) require heterologous expression to realize their genetic potential, including silent and metagenomic BGCs. Although the engineered Streptomyces coelicolor M1152 is a widely used host for heterologous expression of BGCs, a systemic understanding of how its genetic modifications affect the metabolism is lacking and limiting further development. We performed a comparative analysis of M1152 and its ancestor M145, connecting information from proteomics, transcriptomics, and cultivation data into a comprehensive picture of the metabolic differences between these strains. Instrumental to this comparison was the application of an improved consensus genome-scale metabolic model (GEM) of S. coelicolor. Although many metabolic patterns are retained in M1152, we find that this strain suffers from oxidative stress, possibly caused by increased oxidative metabolism. Furthermore, precursor availability is likely not limiting polyketide production, implying that other strategies could be beneficial for further development of S. coelicolor for heterologous production of novel compounds.

scholarly journals Genome mining and biosynthesis of kitacinnamycins as a STING activator

2019 ◽  
Vol 10 (18) ◽  
pp. 4839-4846 ◽  
Author(s):  
Jing Shi ◽  
Cheng Li Liu ◽  
Bo Zhang ◽  
Wen Jie Guo ◽  
Jiapeng Zhu ◽  
...  
Keyword(s):  
Genome Mining ◽  
Type Ii ◽  
Unique Type ◽  
Type Ii Pks

Genome mining targeting unique type II PKS and NRPS led to the identification of a novel class of glycopeptides named kitacinnamycins.

scholarly journals Heterologous Expression of a Cryptic Gene Cluster from Streptomyces leeuwenhoekii C34T Yields a Novel Lasso Peptide, Leepeptin

2019 ◽  
Vol 85 (23) ◽  
Author(s):  
Juan Pablo Gomez-Escribano ◽  
Jean Franco Castro ◽  
Valeria Razmilic ◽  
Scott A. Jarmusch ◽  
Gerhard Saalbach ◽  
...  
Keyword(s):  
Natural Products ◽  
Heterologous Expression ◽  
Streptomyces Coelicolor ◽  
Gene Clusters ◽  
Bioinformatic Analysis ◽  
Growth Media ◽  
Content Type ◽  
Lasso Peptide ◽  
Lasso Peptides ◽  
New Family

ABSTRACT Analysis of the genome sequence of Streptomyces leeuwenhoekii C34T identified biosynthetic gene clusters (BGCs) for three different lasso peptides (Lp1, Lp2, and Lp3) which were not known to be made by the strain. Lasso peptides represent relatively new members of the RiPP (ribosomally synthesized and posttranslationally modified peptides) family of natural products and have not been extensively studied. Lp3, whose production could be detected in culture supernatants from S. leeuwenhoekii C34T and after heterologous expression of its BGC in Streptomyces coelicolor, is identical to the previously characterized chaxapeptin. Lp1, whose production could not be detected or achieved heterologously, appears to be identical to a recently identified member of the citrulassin family of lasso peptides. Since production of Lp2 by S. leeuwenhoekii C34T was not observed, its BGC was also expressed in S. coelicolor. The lasso peptide was isolated and its structure confirmed by mass spectrometry and nuclear magnetic resonance analyses, revealing a novel structure that appears to represent a new family of lasso peptides. IMPORTANCE Recent developments in genome sequencing combined with bioinformatic analysis have revealed that actinomycetes contain a plethora of unexpected BGCs and thus have the potential to produce many more natural products than previously thought. This reflects the inability to detect the production of these compounds under laboratory conditions, perhaps through the use of inappropriate growth media or the absence of the environmental cues required to elicit expression of the corresponding BGCs. One approach to overcoming this problem is to circumvent the regulatory mechanisms that control expression of the BGC in its natural host by deploying heterologous expression. The generally compact nature of lasso peptide BGCs makes them particularly amenable to this approach, and, in the example given here, analysis revealed a new member of the lasso peptide family of RiPPs. This approach should be readily applicable to other cryptic lasso peptide gene clusters and would also facilitate the design and production of nonnatural variants by changing the sequence encoding the core peptide, as has been achieved with other classes of RiPPs.

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