scholarly journals The granaticin biosynthetic gene cluster of Streptomyces violaceoruber Tü22: sequence analysis and expression in a heterologous host

1998 ◽  
Vol 5 (11) ◽  
pp. 647-659 ◽  
Author(s):  
Koji Ichinose ◽  
David J. Bedford ◽  
Diethild Tornus ◽  
Andreas Bechthold ◽  
Maureen J. Bibb ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Gene Cluster ◽  
Biosynthetic Gene Cluster ◽  
Biosynthetic Gene ◽  
Heterologous Host ◽  
Streptomyces Violaceoruber

scholarly journals Molecular Cloning, Sequence Analysis, and Heterologous Expression of the Phosphinothricin Tripeptide Biosynthetic Gene Cluster from Streptomyces viridochromogenes DSM 40736

2005 ◽  
Vol 49 (1) ◽  
pp. 230-240 ◽  
Author(s):  
Joshua A. V. Blodgett ◽  
Jun Kai Zhang ◽  
William W. Metcalf
Keyword(s):  
Sequence Analysis ◽  
Heterologous Expression ◽  
Gene Cluster ◽  
Genomic Dna ◽  
Biosynthetic Gene Cluster ◽  
Fosmid Library ◽  
Open Reading Frames ◽  
Biosynthetic Gene ◽  
Heterologous Host ◽  
Reading Frames

ABSTRACT A fosmid library from genomic DNA of Streptomyces viridochromogenes DSM 40736 was constructed and screened for the presence of genes known to be involved in the biosynthesis of phosphinothricin tripeptide (PTT). Eight positives were identified, one of which was able to confer PTT biosynthetic capability upon Streptomyces lividans after integration of the fosmid into the chromosome of this heterologous host. Sequence analysis of the 40,241-bp fosmid insert revealed 29 complete open reading frames (ORFs). Deletion analysis demonstrated that a minimum set of 24 ORFs were required for PTT production in the heterologous host. Sequence analysis revealed that most of these PTT genes have been previously identified in either S. viridochromogenes or S. hygroscopicus (or both), although only 11 out of 24 of these ORFs have experimentally defined functions. Three previously unknown genes within the cluster were identified and are likely to have roles in the stepwise production of phosphonoformate from phosphonoacetaldehyde. This is the first report detailing the entire PTT gene cluster from any producing streptomycete.

scholarly journals Molecular cloning and sequence analysis of the complestatin biosynthetic gene cluster

2001 ◽  
Vol 98 (15) ◽  
pp. 8548-8553 ◽  
Author(s):  
H.-T. Chiu ◽  
B. K. Hubbard ◽  
A. N. Shah ◽  
J. Eide ◽  
R. A. Fredenburg ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Molecular Cloning ◽  
Gene Cluster ◽  
Biosynthetic Gene Cluster ◽  
Biosynthetic Gene

scholarly journals Identification of a Biosynthetic Gene Cluster Responsible for the Production of a New Pyrrolopyrimidine Natural Product—Huimycin

Biomolecules ◽  
2020 ◽  
Vol 10 (7) ◽  
pp. 1074
Author(s):  
Hui Shuai ◽  
Maksym Myronovskyi ◽  
Suvd Nadmid ◽  
Andriy Luzhetskyy
Keyword(s):  
Gene Cluster ◽  
Gene Deletion ◽  
Biological Activities ◽  
Biosynthetic Gene Cluster ◽  
Resonance Spectroscopy ◽  
Important Class ◽  
Biosynthetic Gene ◽  
Heterologous Host ◽  
Streptomyces Albus ◽  
Actinomycete Strain

Pyrrolopyrimidines are an important class of natural products with a broad spectrum of biological activities, including antibacterial, antifungal, antiviral, anticancer or anti-inflammatory. Here, we present the identification of a biosynthetic gene cluster from the rare actinomycete strain Kutzneria albida DSM 43870, which leads to the production of huimycin, a new member of the pyrrolopyrimidine family of compounds. The huimycin gene cluster was successfully expressed in the heterologous host strain Streptomyces albus Del14. The compound was purified, and its structure was elucidated by means of nuclear magnetic resonance spectroscopy. The minimal huimycin gene cluster was identified through sequence analysis and a series of gene deletion experiments. A model for huimycin biosynthesis is also proposed in this paper.

Analysis of YM-216391 Biosynthetic Gene Cluster and Improvement of the Cyclopeptide Production in a Heterologous Host

2012 ◽  
Vol 7 (4) ◽  
pp. 646-651 ◽  
Author(s):  
Xiao-Hong Jian ◽  
Hai-Xue Pan ◽  
Ting-Ting Ning ◽  
Yuan-Yuan Shi ◽  
Yong-Sheng Chen ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Biosynthetic Gene Cluster ◽  
Biosynthetic Gene ◽  
Heterologous Host

scholarly journals Isolation, Characterization, and Heterologous Expression of the Biosynthesis Gene Cluster for the Antitumor Anthracycline Steffimycin

2006 ◽  
Vol 72 (6) ◽  
pp. 4172-4183 ◽  
Author(s):  
Sonia Gull�n ◽  
Carlos Olano ◽  
Mohamed S. Abdelfattah ◽  
Alfredo F. Bra�a ◽  
J�rgen Rohr ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Biosynthetic Gene Cluster ◽  
Open Reading Frames ◽  
Biosynthetic Gene ◽  
Heterologous Host ◽  
Biosynthesis Gene Cluster ◽  
Polyketide Biosynthesis ◽  
Gene Coding ◽  
Biosynthesis Gene ◽  
Reading Frames

ABSTRACT The biosynthetic gene cluster for the aromatic polyketide steffimycin of the anthracycline family has been cloned and characterized from “Streptomyces steffisburgensis” NRRL 3193. Sequence analysis of a 42.8-kbp DNA region revealed the presence of 36 open reading frames (ORFs) (one of them incomplete), 24 of which, spanning 26.5 kb, are probably involved in steffimycin biosynthesis. They code for all the activities required for polyketide biosynthesis, tailoring, regulation, and resistance but show no evidence of genes involved in l-rhamnose biosynthesis. The involvement of the cluster in steffimycin biosynthesis was confirmed by expression of a region of about 15 kb containing 15 ORFS, 11 of them forming part of the cluster, in the heterologous host Streptomyces albus, allowing the isolation of a biosynthetic intermediate. In addition, the expression in S. albus of the entire cluster, contained in a region of 34.8 kb, combined with the expression of plasmid pRHAM, directing the biosynthesis of l-rhamnose, led to the production of steffimycin. Inactivation of the stfX gene, coding for a putative cyclase, revealed that this enzymatic activity participates in the cyclization of the fourth ring, making the final steps in the biosynthesis of the steffimycin aglycon similar to those in the biosynthesis of jadomycin or rabelomycin. Inactivation of the stfG gene, coding for a putative glycosyltransferase involved in the attachment of l-rhamnose, allowed the production of a new compound corresponding to the steffimycin aglycon compound also observed in S. albus upon expression of the entire cluster.

scholarly journals Heterologous expression of the atypical tetracycline chelocardin reveals the full set of genes required for its biosynthesis

2020 ◽  
Vol 19 (1) ◽  
Author(s):  
Tadeja Lukežič ◽  
Špela Pikl ◽  
Nestor Zaburannyi ◽  
Maja Remškar ◽  
Hrvoje Petković ◽  
...  
Keyword(s):  
Heterologous Expression ◽  
Gene Cluster ◽  
Regulatory Protein ◽  
Model Organism ◽  
Gene Clusters ◽  
Biosynthetic Gene Cluster ◽  
Biosynthetic Gene ◽  
Heterologous Host ◽  
Yield Increase ◽  
Tract Infections

Abstract Background Chelocardin (CHD) exhibits a broad-spectrum antibiotic activity and showed promising results in a small phase II clinical study conducted on patients with urinary tract infections. Importantly, CHD was shown to be active also against tetracycline-resistant Gram-negative pathogens, which is gaining even more importance in today’s antibiotic crisis. We have demonstrated that modifications of CHD through genetic engineering of its producer, the actinomycete Amycolatopsis sulphurea, are not only possible but yielded even more potent antibiotics than CHD itself, like 2-carboxamido-2-deacetyl-chelocardin (CD-CHD), which is currently in preclinical evaluation. A. sulphurea is difficult to genetically manipulate and therefore manipulation of the chd biosynthetic gene cluster in a genetically amenable heterologous host would be of high importance for further drug-discovery efforts. Results We report heterologous expression of the CHD biosynthetic gene cluster in the model organism Streptomyces albus del14 strain. Unexpectedly, we found that the originally defined CHD gene cluster fails to provide all genes required for CHD formation, including an additional cyclase and two regulatory genes. Overexpression of the putative pathway-specific streptomyces antibiotic regulatory protein chdB in A. sulphurea resulted in an increase of both, CHD and CD-CHD production. Applying a metabolic-engineering approach, it was also possible to generate the potent CHD analogue, CD-CHD in S. albus. Finally, an additional yield increase was achieved in S. albus del14 by in-trans overexpression of the chdR exporter gene, which provides resistance to CHD and CDCHD. Conclusions We identified previously unknown genes in the CHD cluster, which were shown to be essential for chelocardin biosynthesis by expression of the full biosynthetic gene cluster in S. albus as heterologous host. When comparing to oxytetracycline biosynthesis, we observed that the CHD gene cluster contains additional enzymes not found in gene clusters encoding the biosynthesis of typical tetracyclines (such as oxytetracycline). This finding probably explains the different chemistries and modes of action, which make CHD/CD-CHD valuable lead structures for clinical candidates. Even though the CHD genes are derived from a rare actinomycete A. sulphurea, the yield of CHD in the heterologous host was very good. The corrected nucleotide sequence of the CHD gene cluster now contains all gene products required for the production of CHD in a genetically amenable heterologous host, thus opening new possibilities towards production of novel and potent tetracycline analogues with a new mode of action.

scholarly journals Cloning and Characterization of the Pyrrolomycin Biosynthetic Gene Clusters from Actinosporangium vitaminophilum ATCC 31673 and Streptomyces sp. Strain UC 11065

2006 ◽  
Vol 51 (3) ◽  
pp. 946-957 ◽  
Author(s):  
Xiujun Zhang ◽  
Ronald J. Parry
Keyword(s):  
Sequence Analysis ◽  
Gene Cluster ◽  
Genetic Locus ◽  
Gene Clusters ◽  
Biosynthetic Gene Cluster ◽  
Open Reading Frames ◽  
Biosynthetic Gene ◽  
Genes Encoding ◽  
Cloned Gene

ABSTRACT The pyrrolomycins are a family of polyketide antibiotics, some of which contain a nitro group. To gain insight into the nitration mechanism associated with the formation of these antibiotics, the pyrrolomycin biosynthetic gene cluster from Actinosporangium vitaminophilum was cloned. Sequencing of ca. 56 kb of A. vitaminophilum DNA revealed 35 open reading frames (ORFs). Sequence analysis revealed a clear relationship between some of these ORFs and the biosynthetic gene cluster for pyoluteorin, a structurally related antibiotic. Since a gene transfer system could not be devised for A. vitaminophilum, additional proof for the identity of the cloned gene cluster was sought by cloning the pyrrolomycin gene cluster from Streptomyces sp. strain UC 11065, a transformable pyrrolomycin producer. Sequencing of ca. 26 kb of UC 11065 DNA revealed the presence of 17 ORFs, 15 of which exhibit strong similarity to ORFs in the A. vitaminophilum cluster as well as a nearly identical organization. Single-crossover disruption of two genes in the UC 11065 cluster abolished pyrrolomycin production in both cases. These results confirm that the genetic locus cloned from UC 11065 is essential for pyrrolomycin production, and they also confirm that the highly similar locus in A. vitaminophilum encodes pyrrolomycin biosynthetic genes. Sequence analysis revealed that both clusters contain genes encoding the two components of an assimilatory nitrate reductase. This finding suggests that nitrite is required for the formation of the nitrated pyrrolomycins. However, sequence analysis did not provide additional insights into the nitration process, suggesting the operation of a novel nitration mechanism.

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