Identification of the genecluster involved in muraymycin biosynthesis from Streptomyces sp. NRRL 30471

Cheng, Lin; Chen, Wenqing; Zhai, Lipeng; Xu, Donghui; Huang, Tingting; Lin, Shuangjun; Zhou, Xing; Deng, Zixin

doi:10.1039/c0mb00237b

Cited by 51 publications

(44 citation statements)

References 29 publications

(56 reference statements)

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“…Identification of the Riboflavin Catabolic Gene Cluster-A cosmid library was constructed for the isolation of the riboflavin catabolic gene cluster (17). After failing to identify the gene cluster by screening this library in Escherichia coli, Streptomyces lividans 1326 was chosen as the heterologous host because Microbacterium and Streptomyces both have similar genomic GC content (ϳ70%).…”

Section: Resultsmentioning

confidence: 99%

Identification of the First Riboflavin Catabolic Gene Cluster Isolated from Microbacterium maritypicum G10

Chakrabarty

Philmus

et al. 2016

Journal of Biological Chemistry

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Riboflavin is a common cofactor, and its biosynthetic pathway is well characterized. However, its catabolic pathway, despite intriguing hints in a few distinct organisms, has never been established. This article describes the isolation of a Microbacterium maritypicum riboflavin catabolic strain, and the cloning of the riboflavin catabolic genes. RcaA, RcaB, RcaD, and RcaE were overexpressed and biochemically characterized as riboflavin kinase, riboflavin reductase, ribokinase, and riboflavin hydrolase, respectively. Based on these activities, a pathway for riboflavin catabolism is proposed.Cofactors play key roles in augmenting the limited functionality available on proteins for catalysis. As with all other metabolites, cofactors do not accumulate in the environment. This is due in part to efficient salvage of these biosynthetically expensive metabolites and in part to cofactor catabolism. However, in contrast to the large literature on cofactor biosynthesis, relatively little is known about cofactor breakdown (1). Detailed enzymology has only been carried out on heme (2), pyridoxal (3), and NAD (4, 5) catabolism. Thiamin, folate, riboflavin, and biotin degrading bacteria were once isolated but have now been lost (6 -11). Analysis of culture metabolites from the reported riboflavin catabolic strain demonstrated that this strain was capable of fully degrading the isoalloxazine ring of riboflavin (12). None of the genes were identified and lumichrome was not an intermediate. Riboflavin hydrolase, the only previously characterized riboflavin catabolic enzyme, catalyzes the conversion of riboflavin to lumichrome and ribose and has previously been identified as a potential catabolic enzyme in a second pathway (13-15). However, despite significant effort, the gene encoding this enzyme was never identified and the enzyme itself proved to be biochemically intractable (16). Here we report the isolation of a riboflavin-catabolizing Microbacterium maritypicum strain from dust samples obtained at the DSM riboflavin production plant in Germany. The catabolic operon was isolated from a cosmid library, sequenced, and annotated. Preliminary characterization of the enzymes involved suggested a riboflavin catabolic pathway. ResultsStrain M. maritypicum G10 Catabolizes Riboflavin-Dust samples, collected at the DSM riboflavin production plant in Germany, were screened by culturing in a defined minimal medium containing M9 salts with riboflavin as the primary carbon source. Bleaching of the riboflavin color was observed in some of the samples. The cells from these hits were grown on NB plates and single colonies were selected. Individual strains were retested in M9 medium with riboflavin as the sole carbon source. The best of these strains (strain G10) completely bleached the yellow color of riboflavin after 12 h of growth and was selected for further study. The sequence of the gene encoding 16S rRNA identified this strain as M. maritypicum.To identify the products generated from consumption of riboflavin, M. maritypicum G10 was gro...

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Section: Resultsmentioning

confidence: 99%

Identification of the First Riboflavin Catabolic Gene Cluster Isolated from Microbacterium maritypicum G10

Chakrabarty

Philmus

et al. 2016

Journal of Biological Chemistry

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“…To achieve engineered production of 2=-Cl PTN and 2=-amino dA in a heterologous host, a positive cosmid, 3G12, housing the probable complete ada gene cluster, was screened from the genomic library of Actinomadura sp. ATCC 39365 by a narrowdown PCR strategy (16) and introduced into the heterologous host Streptomyces aureochromogenes CXR14 (17). Subsequently, the recombinant strain (CXR14::3G12), confirmed by PCR, was fermented for further metabolic analysis, and the LC-MS results indicated that the targeted [MϩH] ϩ ions of 2=-Cl PTN (m/z 303.0825), PTN (m/z 269.1220), and 2=-amino dA (m/z 267.1175) could be clearly detected from the sample of the CXR14::3G12 recombinant.…”

Section: Resultsmentioning

confidence: 99%

Biosynthesis of 2′-Chloropentostatin and 2′-Amino-2′-Deoxyadenosine Highlights a Single Gene Cluster Responsible for Two Independent Pathways in Actinomadura sp. Strain ATCC 39365

Gao

et al. 2017

Appl Environ Microbiol

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2=-Chloropentostatin (2=-Cl PTN, 2=-chloro-2=-deoxycoformycin) and 2=-amino-2=-deoxyadenosine (2=-amino dA) are two adenosine-derived nucleoside antibiotics coproduced by Actinomadura sp. strain ATCC 39365. 2=-Cl PTN is a potent adenosine deaminase (ADA) inhibitor featuring an intriguing 1,3-diazepine ring, as well as a chlorination at C-2= of ribose, and 2=-amino dA is an adenosine analog showing bioactivity against RNA-type virus infection. However, the biosynthetic logic of them has remained poorly understood. Here, we report the identification of a single gene cluster (ada) essential for the biosynthesis of 2=-Cl PTN and 2=-amino dA. Further systematic genetic investigations suggest that 2=-Cl PTN and 2=-amino dA are biosynthesized by independent pathways. Moreover, we provide evidence that a predicted cation/H ϩ antiporter, AdaE, is involved in the chlorination step during 2=-Cl PTN biosynthesis. Notably, we demonstrate that 2=-amino dA biosynthesis is initiated by a Nudix hydrolase, AdaJ, catalyzing the hydrolysis of ATP. Finally, we reveal that the host ADA (designated ADA1), capable of converting adenosine/2=-amino dA to inosine/2=-amino dI, is not very sensitive to the powerful ADA inhibitor pentostatin. These findings provide a basis for the further rational pathway engineering of 2=-Cl PTN and 2=-amino dA production.IMPORTANCE 2=-Cl PTN/PTN and 2=-amino dA have captivated the great interests of scientists, owing to their unusual chemical structures and remarkable bioactivities. However, the precise logic for their biosynthesis has been elusive for decades. Actually, the identification and elucidation of their biosynthetic pathways not only enrich the biochemical repertoire of novel enzymatic reactions but may also lay solid foundations for the pathway engineering and combinatorial biosynthesis of this family of purine nucleoside antibiotics to generate novel hybrid analogs with improved features.KEYWORDS 2=-chloropentostatin, 2=-amino-2=-deoxyadenosine, nucleoside antibiotics, biosynthesis, Nudix hydrolase, adenosine deaminase N ucleoside antibiotics are a large family of important microbial natural products harboring wide-range biological properties and distinctive structural features (1-3). Their biosynthesis generally follows a succinct logic by sequential enzymatic modification of nucleosides or nucleotides originating from primary metabolisms (1). Usually, nucleosides and nucleotides play pleiotropic roles in most fundamental cellular metabolisms; therefore, nucleoside antibiotics are able to target the biosynthesis of diverse biomacromolecules, including nucleic acids, proteins, and glycans (1).

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“…2) (29,30). Additional members of the GlyU-containing nucleoside antibiotics with identified biosynthetic gene clusters include liposidomycins (31), caprazamycins (32), muraymycin (33), muraminomicin (34), and the recently discovered sphaerimicins (35). Thus, the uncovering of these two genes within the clusters for 1 and 2 suggested that the biosynthesis of CarU proceeds via condensation of L-Thr with UA to generate GlyU as a cryptic pathway intermediate.…”

Section: Capuramycin-type Antibiotics Include A-500359s Frommentioning

confidence: 99%

The Biosynthesis of Capuramycin-type Antibiotics

Cai

Goswami

Yang

et al. 2015

Journal of Biological Chemistry

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Background: Several nucleoside antibiotics contain a uridine-5Ј-carboxamide core of unclear origin. Results: The A-102395 biosynthetic gene cluster was cloned, a genetic system was developed, and three enzymes were characterized in vivo and in vitro. Conclusion: Uridine-5Ј-carboxamide originates from UMP and L-Thr by sequential reactions catalyzed by a dioxygenase and transaldolase. Significance: The results provide the first opportunity to methodically interrogate the biosynthesis of these unusual antibiotics.

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Identification of the genecluster involved in muraymycin biosynthesis from Streptomyces sp. NRRL 30471

Cited by 51 publications

References 29 publications

Identification of the First Riboflavin Catabolic Gene Cluster Isolated from Microbacterium maritypicum G10

Identification of the First Riboflavin Catabolic Gene Cluster Isolated from Microbacterium maritypicum G10

Biosynthesis of 2′-Chloropentostatin and 2′-Amino-2′-Deoxyadenosine Highlights a Single Gene Cluster Responsible for Two Independent Pathways in Actinomadura sp. Strain ATCC 39365

The Biosynthesis of Capuramycin-type Antibiotics

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