The bengamides, sponge‐derived natural products that have been characterized as inhibitors of methionine aminopeptidases (MetAPs), have been intensively investigated as anticancer compounds. We ...embarked on a multidisciplinary project to supply bengamides by fermentation of the terrestrial myxobacterium M. virescens, decipher their biosynthesis, and optimize their properties as drug leads. The characterization of the biosynthetic pathway revealed that bacterial resistance to bengamides is conferred by Leu 154 of the myxobacterial MetAP protein, and enabled transfer of the entire gene cluster into the more suitable production host M. xanthus DK1622. A combination of semisynthesis of microbially derived bengamides and total synthesis resulted in an optimized derivative that combined high cellular potency in the nanomolar range with high metabolic stability, which translated to an improved half‐life in mice and antitumor efficacy in a melanoma mouse model.
A terrestrial dive to bengamides: Bengamides, sponge‐derived natural products, have been generated from a terrestrial source. Their biosynthesis and self‐resistance mechanism against methionine aminopeptidases was elucidated, a heterologous expression platform was established, and their pharmaceutical properties were improved by medicinal chemistry.
Kendomycin is a bioactive polyketide that is produced by various Streptomyces strains. It displays strong antibiotic activities against a wide range of bacteria and exhibits remarkable cytotoxic ...effects on the growth of several human cancer cell lines. In this study we cloned the corresponding biosynthetic locus from the producer Streptomyces violaceoruber (strain 3844-33C). Our analysis shows that a mixed type I/type III polyketide synthase pathway is responsible for the formation of the fully carbogenic macrocyclic scaffold of kendomycin, which is unprecedented among all of the ansa compounds that have been isolated so far. Heterologous expression of a gene set in Streptomyces coelicolor shows that 3,5-dihydroxybenzoic acid is an intermediate in the starter unit biosynthesis that is initiated by the type III polyketide synthase. The identification of the kendomycin biosynthetic gene cluster sets the stage to study a novel chain termination mechanism by a type I PKS that leads to carbocycle formation and provides the starting material for the heterologous expression of the entire pathway, and the production of novel derivatives by genetic engineering.
Sorangium cellulosum strains produce approximately 50% of the biologically active secondary metabolites known from myxobacteria. These metabolites include several compounds of biotechnological ...importance such as the epothilones and chivosazols, which, respectively, stabilize the tubulin and actin skeletons of eukaryotic cells. S. cellulosum is characterized by its slow growth rate, and natural products are typically produced in low yield. In this study, biomagnetic bead separation of promoter-binding proteins and subsequent inactivation experiments were employed to identify the chivosazol regulator, ChiR, as a positive regulator of chivosazol biosynthesis in the genome-sequenced strain So ce56. Overexpression of chiR under the control of T7A1 promoter in a merodiploid mutant resulted in fivefold overproduction of chivosazol in a kinetic shake flask experiment, and 2.5-fold overproduction by fermentation. Using quantitative reverse transcription PCR and gel shift experiments employing heterologously expressed ChiR, we have shown that transcription of the chivosazol biosynthetic genes (chiA-chiF) is directly controlled by this protein. In addition, we have demonstrated that ChiR serves as a pleiotropic regulator in S. cellulosum, because mutant strains lack the ability to develop into regular fruiting bodies.
Sorangium cellulosum strains produce approximately 50% of the biologically active secondary metabolites known from myxobacteria. These metabolites include several compounds of biotechnological ...importance such as the epothilones and chivosazols, which, respectively, stabilize the tubulin and actin skeletons of eukaryotic cells. S. cellulosum is characterized by its slow growth rate, and natural products are typically produced in low yield. In this study, biomagnetic bead separation of promoter-binding proteins and subsequent inactivation experiments were employed to identify the chivosazol regulator, ChiR, as a positive regulator of chivosazol biosynthesis in the genome-sequenced strain So ceS6. Overexpression of chiR under the control of T7A1 promoter in a merodiploid mutant resulted in fivefold overproduction of chivosazol in a kinetic shake flask experiment, and 2.5-fold overproduction by fermentation. Using quantitative reverse transcription PCR and gel shift experiments employing heterologously expressed ChiR, we have shown that transcription of the chivosazol biosynthetic genes (chiA-chiF) is directly controlled by this protein. In addition, we have demonstrated that ChiR serves as a pleiotropic regulator in S. cellulosum, because mutant strains lack the ability to develop into regular fruiting bodies. PUBLICATION ABSTRACT
The jaspamide/chondramide family of depsipeptides are mixed PKS/NRPS natural products isolated from marine sponges and a terrestrial myxobacterium that potently affect the function of the actin ...cytoskeleton. As a first step to improve production in heterologous host cells and permit genetic approaches to novel analogs, we have cloned and characterized the chondramide biosynthetic genes from the myxobacterium
Chondromyces crocatus Cm c5. In addition to the expected PKS and NRPS genes, the cluster encodes a rare tyrosine aminomutase for β-tyrosine formation and a previously unknown tryptophan-2-halogenase. Conditions for gene transfer into
C. crocatus Cm c5 were developed, and inactivation of several genes corroborated their proposed function and served to define the boundaries of the cluster. Biochemical characterization of the final NRPS adenylation domain confirmed the direct activation of β-tyrosine, and fluorinated chondramides were produced through precursor-directed biosynthesis.
Bengamide, aus marinen Schwämmen stammende und als Inhibitoren der Methionin‐Aminopeptidase (MetAP) charakterisierte Naturstoffe, wurden intensiv als Wirkstoffe gegen Krebs erforscht. In einem ...multidisziplinären Forschungsprojekt haben wir die Bereitstellung von Bengamiden über Fermentation des terrestrischen Myxobakteriums M. virescens, die Aufklärung ihrer Biosynthese und die Optimierung ihrer Eigenschaften als Arzneistoff‐Leitstrukturen untersucht. Die Charakterisierung des Biosyntheseweges zeigte auf, dass bakterielle Resistenz gegenüber Bengamiden durch Leu154 des myxobakteriellen MetAP‐Proteins vermittelt wird, und ermöglichte den Transfer des gesamten Biosynthesegenclusters in den geeigneteren Produktionsstamm M. xanthus DK1622. Eine Kombination aus Semisynthese mikrobiell gewonnener Bengamide und Totalsynthese führte zum optimierten Derivat 8 a. Die nanomolare zelluläre Wirksamkeit und die hohe metabolische Stabilität waren mit einer verbesserten Halbwertszeit in Mäusen sowie mit Antitumor‐Effizienz in einem Melanom‐Mausmodell verbunden.
Ein terrestrischer Tauchgang zu Bengamiden: Bengamide, aus marinen Schwämmen stammende Naturstoffe, wurden in einem terrestrischen Produzenten hergestellt. Ihre Biosynthese und Selbstresistenzmechanismen gegen Methionin‐Aminopeptidasen wurden aufgeklärt, eine heterologe Expressionsplattform wurde etabliert und auf ihre pharmakologischen Eigenschaften optimiert.
Abstract
Bengamide, aus marinen Schwämmen stammende und als Inhibitoren der Methionin‐Aminopeptidase (MetAP) charakterisierte Naturstoffe, wurden intensiv als Wirkstoffe gegen Krebs erforscht. In ...einem multidisziplinären Forschungsprojekt haben wir die Bereitstellung von Bengamiden über Fermentation des terrestrischen Myxobakteriums M. virescens, die Aufklärung ihrer Biosynthese und die Optimierung ihrer Eigenschaften als Arzneistoff‐Leitstrukturen untersucht. Die Charakterisierung des Biosyntheseweges zeigte auf, dass bakterielle Resistenz gegenüber Bengamiden durch Leu154 des myxobakteriellen MetAP‐Proteins vermittelt wird, und ermöglichte den Transfer des gesamten Biosynthesegenclusters in den geeigneteren Produktionsstamm M. xanthus DK1622. Eine Kombination aus Semisynthese mikrobiell gewonnener Bengamide und Totalsynthese führte zum optimierten Derivat
8 a
. Die nanomolare zelluläre Wirksamkeit und die hohe metabolische Stabilität waren mit einer verbesserten Halbwertszeit in Mäusen sowie mit Antitumor‐Effizienz in einem Melanom‐Mausmodell verbunden.
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