Discovery and overproduction of novel highly bioactive pamamycins through transcriptional engineering of the biosynthetic gene cluster

Background Pamamycins are a family of highly bioactive macrodiolide polyketides produced by Streptomyces alboniger as a complex mixture of derivatives with molecular weights ranging from 579 to 705 Daltons. The large derivatives are produced as a minor fraction, which has prevented their isolation and thus studies of chemical and biological properties. Results Herein, we describe the transcriptional engineering of the pamamycin biosynthetic gene cluster (pam BGC), which resulted in the shift in production profle toward high molecular weight derivatives. The pam BGC library was constructed by inserting randomized promoter sequences in front of key biosynthetic operons. The library was expressed in Streptomyces albus strain with improved resistance to pamamycins to overcome sensitivity-related host limitations. Clones with modifed pamamycin profles were selected and the properties of engineered pam BGC were studied in detail. The production level and composition of the mixture of pamamycins was found to depend on balance in expression of the corresponding biosynthetic genes. This approach enabled the isolation of known pamamycins and the discovery of three novel derivatives with molecular weights of 663 Da and higher. One of them, homopamamycin 677A, is the largest described representative of this family of natural products with an elucidated structure. The new pamamycin 663A shows extraordinary activity (IC50 2 nM) against hepatocyte cancer cells as well as strong activity (in the one-digit micromolar range) against a range of Gram-positive pathogenic bacteria. Conclusion By employing transcriptional gene cluster refactoring, we not only enhanced the production of known pamamycins but also discovered novel derivatives exhibiting promising biological activities. This approach has the potential for broader application in various biosynthetic gene clusters, creating a sustainable supply and discovery platform for bioactive natural products

Prefrontal and anterior cingulate cortex abnormalities in Tourette Syndrome: evidence from voxel-based morphometry and magnetization transfer imaging

<p>Abstract</p> <p>Background</p> <p>Pathophysiological evidence suggests an involvement of fronto-striatal circuits in Tourette syndrome (TS). To identify TS related abnormalities in gray and white matter we used optimized voxel-based morphometry (VBM) and magnetization transfer imaging (MTI) which are more sensitive to tissue alterations than conventional MRI and provide a quantitative measure of macrostructural integrity.</p> <p>Methods</p> <p>Volumetric high-resolution anatomical T1-weighted MRI and MTI were acquired in 19 adult, unmedicated male TS patients without co-morbidities and 20 age- and sex-matched controls on a 1.5 Tesla neuro-optimized GE scanner. Images were pre-processed and analyzed using an optimized version of VBM in SPM2.</p> <p>Results</p> <p>Using VBM, TS patients showed significant decreases in gray matter volumes in prefrontal areas, the anterior cingulate gyrus, sensorimotor areas, left caudate nucleus and left postcentral gyrus. Decreases in white matter volumes were detected in the right inferior frontal gyrus, the left superior frontal gyrus and the anterior corpus callosum. Increases were found in the left middle frontal gyrus and left sensorimotor areas. In MTI, white matter reductions were seen in the right medial frontal gyrus, the inferior frontal gyrus bilaterally and the right cingulate gyrus. Tic severity was negatively correlated with orbitofrontal structures, the right cingulate gyrus and parts of the parietal-temporal-occipital association cortex bilaterally.</p> <p>Conclusion</p> <p>Our MRI <it>in vivo </it>neuropathological findings using two sensitive and unbiased techniques support the hypothesis that alterations in frontostriatal circuitries underlie TS pathology. We suggest that anomalous frontal lobe association and projection fiber bundles cause disinhibition of the cingulate gyrus and abnormal basal ganglia function.</p

Untersuchungen zur Schallimmissionsprognose bei tieffrequenten Geräuschen

In der Veröffentlichung werden Messungen der Schallübertragung bei tiefen Frequenzen an Gebäuden in Sachsen beschrieben. Die Ergebnisse ermöglichen es, bei der Einwirkung tieffrequenter Geräusche von außen den Schalldruckpegel im Raum abzuschätzen. Anhand einer Literaturrecherche wird der gegenwärtige Erkenntnisstand erörtert. Theoretische Betrachtungen und Simulationsrechnungen ergänzen die Messergebnisse. Die Veröffentlichung richtet sich an Ingenieure, Planer und Immissionsschutzbehörden. Redaktionsschluss: 31.01.202

Untersuchungen zur Schallimmissionsprognose bei tieffrequenten Geräuschen

In der Veröffentlichung werden Messungen der Schallübertragung bei tiefen Frequenzen an Gebäuden in Sachsen beschrieben. Die Ergebnisse ermöglichen es, bei der Einwirkung tieffrequenter Geräusche von außen den Schalldruckpegel im Raum abzuschätzen. Anhand einer Literaturrecherche wird der gegenwärtige Erkenntnisstand erörtert. Theoretische Betrachtungen und Simulationsrechnungen ergänzen die Messergebnisse. Die Veröffentlichung richtet sich an Ingenieure, Planer und Immissionsschutzbehörden. Redaktionsschluss: 31.01.202

Discovery and overproduction of novel highly bioactive pamamycins through transcriptional engineering of the biosynthetic gene cluster

Eckert N, Rebets Y, Horbal L, et al. Discovery and overproduction of novel highly bioactive pamamycins through transcriptional engineering of the biosynthetic gene cluster. Microbial Cell Factories . 2023;22(1): 233.BACKGROUND: Pamamycins are a family of highly bioactive macrodiolide polyketides produced by Streptomyces alboniger as a complex mixture of derivatives with molecular weights ranging from 579 to 705 Daltons. The large derivatives are produced as a minor fraction, which has prevented their isolation and thus studies of chemical and biological properties.; RESULTS: Herein, we describe the transcriptional engineering of the pamamycin biosynthetic gene cluster (pam BGC), which resulted in the shift in production profile toward high molecular weight derivatives. The pam BGC library was constructed by inserting randomized promoter sequences in front of key biosynthetic operons. The library was expressed in Streptomyces albus strain with improved resistance to pamamycins to overcome sensitivity-related host limitations. Clones with modified pamamycin profiles were selected and the properties of engineered pam BGC were studied in detail. The production level and composition of the mixture of pamamycins was found to depend on balance in expression of the corresponding biosynthetic genes. This approach enabled the isolation of known pamamycins and the discovery of three novel derivatives with molecular weights of 663 Da and higher. One of them, homopamamycin 677A, is the largest described representative of this family of natural products with an elucidated structure. The new pamamycin 663A shows extraordinary activity (IC50 2nM) against hepatocyte cancer cells as well as strong activity (in the one-digit micromolar range) against a range of Gram-positive pathogenic bacteria.; CONCLUSION: By employing transcriptional gene cluster refactoring, we not only enhanced the production of known pamamycins but also discovered novel derivatives exhibiting promising biological activities. This approach has the potential for broader application in various biosynthetic gene clusters, creating a sustainable supply and discovery platform for bioactive natural products. © 2023. The Author(s)