Complete Genome Sequence of Actinosynnema pretiosum X47, An Industrial Strain that Produces the Antibiotic Ansamitocin AP-3

Zhong, Chunlong; Zong, Guanghua; Qian, Shulan; Li, Meng; Fu, Jiafang; Zhang, Peipei; Li, Jun; Cao, Guangxiang

doi:10.1007/s00284-018-1521-1

Cited by 6 publications

(9 citation statements)

References 20 publications

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“…generated a high AP-3 yield A . pretiosum ATCC 31565, in which the production of AP-3 increased up to 19.7-fold [ 18 ]. Although mutagenesis has shown advantages in obtaining new strains, completely random mutations in the host genome critically dampen the subsequent screening efficiency.…”

Section: Discussionmentioning

confidence: 99%

“…Traditional mutagenesis and specific gene modification represent two major ways to obtain AP-3 high-yield strains. By traditional methods, high-yield strains can be obtained by physical irradiation- and compound-induced mutagenesis [ 17 , 18 ]. However, mutations are usually accompanied by unexpected randomness, which makes it difficult to achieve precise control of the direction of evolution.…”

Section: Introductionmentioning

confidence: 99%

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Increased yield of AP-3 by inactivation of asm25 in Actinosynnema pretiosum ssp. auranticum ATCC 31565

Cheng

Xiong

et al. 2022

PLoS ONE

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Asamitocins are maytansinoids produced by Actinosynnema pretiosum ssp. auranticum ATCC 31565 (A. pretiosum ATCC 31565), which have a structure similar to that of maytansine, therefore serving as a precursor of maytansine in the development of antibody-drug conjugates (ADCs). Currently, there are more than 20 known derivatives of ansamitocins, among which ansamitocin P-3 (AP-3) exhibits the highest antitumor activity. Despite its importance, the application of AP-3 is restricted by low yield, likely due to a substrate competition mechanism underlying the synthesis pathways of AP-3 and its byproducts. Given that N-demethylansamitocin P-3, the precursor of AP-3, is regulated by asm25 and asm10 to synthesize AGP-3 and AP-3, respectively, asm25 is predicted to be an inhibitory gene for AP-3 production. In this study, we inactivated asm25 in A. pretiosum ATCC 31565 by CRISPR-Cas9-guided gene editing. asm25 depletion resulted in a more than 2-fold increase in AP-3 yield. Surprisingly, the addition of isobutanol further improved AP-3 yield in the asm25 knockout strain by more than 6 times; in contrast, only a 1.53-fold increase was found in the WT strain under the parallel condition. Thus, we uncovered an unknown function of asm25 in AP-3 yield and identified asm25 as a promising target to enhance the large-scale industrial production of AP-3.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Increased yield of AP-3 by inactivation of asm25 in Actinosynnema pretiosum ssp. auranticum ATCC 31565

Cheng

Xiong

et al. 2022

PLoS ONE

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“…Actinosynnema pretiosum X47 in this study was derived from A. pretiosum subspecies auranticum ATCC 31565. The genome of A. pretiosum X47 was about 8.13 Mb in length, with an average GC content of 73.91%, and 7029 genes were predicted, including 45 pairs of putative two-component signal transduction system (TCS), 13 histidine kinases, and 38 orphan response regulators ( Zhong et al, 2019 ). The morphological development of A. pretiosum is complex and similar to that of Streptomyces .…”

Section: Introductionmentioning

confidence: 99%

“…Although PhoPR is essential for the normal growth and metabolism of a broad range of species, the regulatory functions and mechanisms of PhoP differ in different bacteria. In this study, the PhoPR homolog was identified in the genome of the A. pretiosum X47 strain ( Zhong et al, 2019 ), and a phoP mutant strain was constructed. Our data suggest that deletion of phoP accelerates the formation of aerial mycelia and spore formation in strain X47, contributing to a better understanding of developmental regulation in A. pretiosum strains.…”

Section: Introductionmentioning

confidence: 99%

Deletion of the Response Regulator PhoP Accelerates the Formation of Aerial Mycelium and Spores in Actinosynnema pretiosum

Zhang

Liu

et al. 2022

Front. Microbiol.

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PhoPR is an important two-component signal transduction system (TCS) for microorganisms to sense and respond to phosphate limitation. Although the response regulator PhoP controls morphological development and secondary metabolism in various Streptomyces species, the function of PhoP in Actinosynnema pretiosum remains unclear. In this study, we showed that PhoP significantly represses the morphological development of the A. pretiosum X47 strain. Production of aerial mycelium and spore formation occurred much earlier in the ΔphoP strain than in X47 during growth on ISP2 medium. Transcription analysis indicated that 222 genes were differentially expressed in ∆phoP compared to strain X47. Chemotaxis genes (cheA, cheW, cheX, and cheY); flagellum biosynthesis and motility genes (flgBCDGKLN, flaD, fliD-R, motA, and swrD); and differentiation genes (whiB and ssgB) were significantly upregulated in ∆phoP. Gel-shift analysis indicated that PhoP binds to the promoters of flgB, flaD, and ssgB genes, and PHO box-like motif with the 8-bp conserved sequence GTTCACGC was identified. The transcription of phoP/phoR of X47 strain was induced at low phosphate concentration. Our results demonstrate that PhoP is a negative regulator that controls the morphological development of A. pretiosum X47 by repressing the transcription of differentiation genes.

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“…Although species in this genus are limited, Actinosynnema -origined natural products exhibit a number of biological and pharmacological activities, such as antitumor (ansamitocin [ 4 , 5 , 6 , 7 ], dnacin B1 [ 8 ] and actinosynneptides [ 9 ]), antibacterial (nocardicins [ 10 ]), anti-fungi (validoxylamine A [ 11 ]), metalloprotease inhibitor (propioxatin [ 12 ]), and siderophore (mirubactin [ 13 ]) ( Figure 1 A). Its promising application in pharmacy promoted the sequencing of the completed genome of four Actinosynnema strains (including two type strains) [ 14 , 15 , 16 , 17 ], which not only boosted the titer optimization of bioactive products by systematic metabolic engineering, but also facilitated the discovery of the Actinosynnema -derived antibiotics and dissection of their biosynthetic machineries.…”

Section: Introductionmentioning

confidence: 99%

p-Aminophenylalanine Involved in the Biosynthesis of Antitumor Dnacin B1 for Quinone Moiety Formation

Xing

Sheng

et al. 2020

Molecules

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Actinosynnema species produce diverse natural products with important biological activities, which represent an important resource of antibiotic discovery. Advances in genome sequencing and bioinformatics tools have accelerated the exploration of the biosynthetic gene clusters (BGCs) encoding natural products. Herein, the completed BGCs of dnacin B1 were first discovered in two Actinosynnema pretiosum subsp. auranticum strains DSM 44131T (hereafter abbreviated as strain DSM 44131T) and X47 by comparative genome mining strategy. The BGC for dnacin B1 contains 41 ORFs and spans a 66.9 kb DNA region in strain DSM 44131T. Its involvement in dnacin B1 biosynthesis was identified through the deletion of a 9.7 kb region. Based on the functional gene analysis, we proposed the biosynthetic pathway for dnacin B1. Moreover, p-amino-phenylalanine (PAPA) unit was found to be the dnacin B1 precursor for the quinone moiety formation, and this was confirmed by heterologous expression of dinV, dinE and dinF in Escherichia coli. Furthermore, nine potential PAPA aminotransferases (APAT) from the genome of strain DSM 44131T were explored and expressed. Biochemical evaluation of their amino group transformation ability was carried out with p-amino-phenylpyruvic acid (PAPP) or PAPA as the substrate for the final product formation. Two of those, APAT4 and APAT9, displayed intriguing aminotransferase ability for the formation of PAPA. The proposed dnacin B1 biosynthetic machinery and PAPA biosynthetic investigations not only enriched the knowledge of tetrahydroisoquinoline (THIQ) biosynthesis, but also provided PAPA building blocks to generate their structurally unique homologues.

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Complete Genome Sequence of Actinosynnema pretiosum X47, An Industrial Strain that Produces the Antibiotic Ansamitocin AP-3

Cited by 6 publications

References 20 publications

Increased yield of AP-3 by inactivation of asm25 in Actinosynnema pretiosum ssp. auranticum ATCC 31565

Increased yield of AP-3 by inactivation of asm25 in Actinosynnema pretiosum ssp. auranticum ATCC 31565

Deletion of the Response Regulator PhoP Accelerates the Formation of Aerial Mycelium and Spores in Actinosynnema pretiosum

p-Aminophenylalanine Involved in the Biosynthesis of Antitumor Dnacin B1 for Quinone Moiety Formation

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