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Sideromycins are a unique subset of siderophores comprising of a siderophore conjugated to an antimicrobial agent. The "Trojan horse" antibiotic albomycins are unique sideromycins consisting of a ferrichrome-type siderophore conjugated to a peptidyl nucleoside antibiotic. They exhibit potent antibacterial activities against many model bacteria and a number of clinical pathogens. Earlier studies have provided significant insight into the biosynthetic pathway of the peptidyl nucleoside moiety. We herein decipher the biosynthetic pathway of the ferrichrome-type siderophore in Streptomyces sp. ATCC 700974. Our genetic studies suggested that abmA, abmB, and abmQ are involved in the formation of the ferrichrome-type siderophore. Additionally, we performed biochemical studies to demonstrate that a flavin-dependent monooxygenase AbmB and an N-acyltransferase AbmA catalyze sequential modifications of L-ornithine to generate N5-acetyl-N5-hydroxyornithine. Three molecules of N5-acetyl-N5-hydroxyornithine are then assembled to generate the tripeptide ferrichrome through the action of a nonribosomal peptide synthetase AbmQ. Of special note, we found out that orf05026 and orf03299, two genes scattered elsewhere in the chromosome of Streptomyces sp. ATCC 700974, have functional redundancy for abmA and abmB, respectively. Interestingly, both orf05026 and orf03299 are situated within gene clusters encoding putative siderophores. In summary, this study provided new insight into the siderophore moiety of albomycin biosynthesis and shed light on the contingency of multiple siderophores in albomycin-producing Streptomyces sp. ATCC 700974.
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Sideróforos , Streptomyces , Sideróforos/metabolismo , Ferricromo/química , Ferricromo/metabolismo , Streptomyces/genética , Streptomyces/metabolismo , Vias Biossintéticas , Nucleosídeos/metabolismo , Antibacterianos/metabolismoRESUMO
Streptomyces bingchenggensis is an industrial producer of milbemycins, which are important anthelmintic and insecticidal agents. Two-component systems (TCSs), which are typically situated in the same operon and are composed of a histidine kinase and a response regulator, are the predominant signal transduction pathways involved in the regulation of secondary metabolism in Streptomyces. Here, an atypical TCS, AtcR/AtcK, in which the encoding genes (sbi_06838/sbi_06839) are organized in a head-to-head pair, was demonstrated to be indispensable for the biosynthesis of multiple secondary metabolites in S. bingchenggensis. With the null TCS mutants, the production of milbemycin and yellow compound was abolished but nanchangmycin was overproduced. Transcriptional analysis and electrophoretic mobility shift assays showed that AtcR regulated the biosynthesis of these three secondary metabolites by a MilR3-mediated cascade. First, AtcR was activated by phosphorylation from signal-triggered AtcK. Second, the activated AtcR promoted the transcription of milR3. Third, MilR3 specifically activated the transcription of downstream genes from milbemycin and yellow compound biosynthetic gene clusters (BGCs) and nanR4 from the nanchangmycin BGC. Finally, because NanR4 is a specific repressor in the nanchangmycin BGC, activation of MilR3 downstream genes led to the production of yellow compound and milbemycin but inhibited nanchangmycin production. By rewiring the regulatory cascade, two strains were obtained, the yield of nanchangmycin was improved by 45-fold to 6.08 g/L and the production of milbemycin was increased twofold to 1.34 g/L. This work has broadened our knowledge on atypical TCSs and provided practical strategies to engineer strains for the production of secondary metabolites in Streptomyces.IMPORTANCEStreptomyces bingchenggensis is an important industrial strain that produces milbemycins. Two-component systems (TCSs), which consist of a histidine kinase and a response regulator, are the predominant signal transduction pathways involved in the regulation of secondary metabolism in Streptomyces. Coupled encoding genes of TCSs are typically situated in the same operon. Here, TCSs with encoding genes situated in separate head-to-head neighbor operons were labeled atypical TCSs. It was found that the atypical TCS AtcR/AtcK played an indispensable role in the biosynthesis of milbemycin, yellow compound, and nanchangmycin in S. bingchenggensis. This atypical TCS regulated the biosynthesis of specialized metabolites in a cascade mediated via a cluster-situated regulator, MilR3. Through rewiring the regulatory pathways, strains were successfully engineered to overproduce milbemycin and nanchangmycin. To the best of our knowledge, this is the first report on atypical TCS, in which the encoding genes of RR and HK were situated in separate head-to-head neighbor operons, involved in secondary metabolism. In addition, data mining showed that atypical TCSs were widely distributed in actinobacteria.
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Éteres , Macrolídeos , Compostos de Espiro , Streptomyces , Histidina Quinase/metabolismo , Streptomyces/genética , Proteínas de Bactérias/genéticaRESUMO
BACKGROUND: Human lysozyme (hLYZ) is a natural antibacterial protein with broad applications in food and pharmaceutical industries. Recombinant production of hLYZ in Komagataella phaffii (K. phaffii) has attracted considerable attention, but there are very limited strategies for its hyper-production in yeast. RESULTS: Here through Atmospheric and Room Temperature Plasma (ARTP)-based mutagenesis and transcriptomic analysis, the expression of two genes MYO1 and IQG1 encoding the cytokinesis core proteins was identified downregulated along with higher hLYZ production. Deletion of either gene caused severe cytokinesis defects, but significantly enhanced hLYZ production. The highest hLYZ yield of 1,052,444 ± 23,667 U/mL bioactivity and 4.12 ± 0.11 g/L total protein concentration were obtained after high-density fed-batch fermentation in the Δmyo1 mutant, representing the best production of hLYZ in yeast. Furthermore, O-linked mannose glycans were characterized on this recombinant hLYZ. CONCLUSIONS: Our work suggests that cytokinesis-based morphology engineering is an effective way to enhance the production of hLYZ in K. phaffii.
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Muramidase , Proteínas Recombinantes , Saccharomycetales , Muramidase/metabolismo , Muramidase/genética , Muramidase/biossíntese , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Saccharomycetales/metabolismo , Saccharomycetales/genética , Humanos , Fermentação , Citocinese , Engenharia Metabólica/métodos , Técnicas de Cultura Celular por LotesRESUMO
Because of the outstanding contribution in genome editing, CRISPR has undoubtedly become the most popular technology around the world and two pioneers are awarded the Nobel Prize in Chemistry this year. Besides, along with the discovery of nonspecific trans-cleavage activities of several Cas proteins such as Cas12 and Cas13, many CRISPR-based molecular diagnostic systems have been successfully created, showing advantages in sensitivity, specificity and operation convenience. Among them, systems with Cas12, which targets DNA and trans-cleaves single-stranded DNA probes, are both simple and highly efficient. Here in this review, we mainly focus on the Cas12-based methods and briefly discuss their applications in nucleic acids detection and beyond.
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Sistemas CRISPR-Cas , Edição de Genes , Sistemas CRISPR-Cas/genética , DNA/genética , DNA de Cadeia Simples , Edição de Genes/métodosRESUMO
Milbemycins (MILs), a group of 16-membered insecticidal macrocylic lactones, are widely used as the biological pesticide and the precursors of semi-synthetic veterinary drugs. Polyketide synthases (PKSs), which require phosphopantetheinyl transferases (PPTases) to activate their ACP domains from apo forms to holo forms, catalyze the backbone biosynthesis of MILs. Here we found there was a complex phosphopantetheinylation network mediated by five putative PPTases in Streptomyces bingchenggensis. Repression mutants of PpA27 and PpA62 via CRISPRi both produced significantly lower yields of MILs than that of the control strain. Repression mutant of PpA68 led to abolishment of the pigment production. MILs production was significantly enhanced by PpA27 overexpression, while not by the overexpression of other PPTases. PpA27 was thus proved a dedicated post-translational enzyme to activate PKSs involved in the MILs biosynthesis. MILs titer was further enhanced by co-overexpression of PpA27 and MilR, the pathwayspecific transcriptional activator of MIL biosynthetic gene cluster. When PpA27 and MilR were co-overexpressed in the industrial S. bingchenggensis HMB, MILs production was increased by 40.5%. These results indicated that tuning the antibiotic biosynthetic pathway by co-engineering transcriptional regulation network and post-translational phosphopantetheinylation network is an effective strategy for antibiotic production improvement.
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Antibacterianos , Macrolídeos , Macrolídeos/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Policetídeo Sintases/genéticaRESUMO
The Great Resignation has brought significant challenges to the recovery of the hospitality industry from the depression caused by the coronavirus pandemic (COVID-19). Prior studies have revealed that the leading cause of the Great Resignation is negative employee experience. However, few empirical studies have been conducted to obtain deep insights into the negative experiences of hospitality employees. Hotel managers still lack the knowledge to help them resolve the workforce problem and maintain competitiveness during the pandemic. This study proposes a novel framework, named HENEX, that uses data-mining technologies and employees' online reviews about hotels to identify the factors that lead to hospitality employees' negative experiences and changes in these factors caused by COVID-19. We demonstrate the effectiveness of HENEX through a case study that involves major hotels in Australia. The findings could help hotel managers develop strategies to resolve the workforce problem and maintain competitiveness during the Great Resignation period.
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Streptomyces bingchenggensis is the main industrial producer of milbemycins, which are a group of 16-membered macrocylic lactones with excellent insecticidal activities. In the past several decades, scientists have made great efforts to solve its low productivity. However, a lack of understanding of the regulatory network of milbemycin biosynthesis limited the development of high-producing strains using a regulatory rewiring strategy. SARPs (Streptomyces Antibiotic Regulatory Proteins) family regulators are widely distributed and play key roles in regulating antibiotics production in actinobacteria. In this paper, MilR3 (encoded by sbi_06842) has been screened out for significantly affecting milbemycin production from all the 19 putative SARP family regulators in S. bingchenggensis with the DNase-deactivated Cpf1-based integrative CRISPRi system. Interestingly, milR3 is about 7 Mb away from milbemycin biosynthetic gene cluster and adjacent to a putative type II PKS (the core minimal PKS encoding genes are sbi_06843, sbi_06844, sbi_06845 and sbi_06846) gene cluster, which was proved to be responsible for producing a yellow pigment. The quantitative real-time PCR analysis proved that MilR3 positively affected the transcription of specific genes within milbemycin BGC and those from the type II PKS gene cluster. Unlike previous "small" SARP family regulators that played pathway-specific roles, MilR3 was probably a unique SARP family regulator and played a pleotropic role. MilR3 was an upper level regulator in the MilR3-MilR regulatory cascade. This study first illustrated the co-regulatory role of this unique SARP regulator. This greatly enriches our understanding of SARPs and lay a solid foundation for milbemycin yield enhancement in the near future.
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Regulação Bacteriana da Expressão Gênica , Streptomyces , Antibacterianos/metabolismo , Desoxirribonucleases/genética , Streptomyces/genética , Streptomyces/metabolismoRESUMO
BACKGROUND: Colletotrichum gloeosporioides ES026, isolated as an endophytic fungal strain, was found to produce the important medicinal compound HuperzineA (HupA). In a genetic context, ES026 showed potential in elucidating the biosynthetic pathway of HupA. METHODS AND RESULTS: The ES026 strain was sequenced using de-novo Illumina sequencing methods in this study. Assembling the cleaned data resulted in 58,594,804bp, consisting of 404 scaffolds. The G + C mol % content of this genome was 52.53%. The genome progressive-alignment with other 4 Colletotrichum strains revealed that ES026 showed closer relation with 030206, SMCG1#C and Nara gc5. More than 60 putative biosynthetic clusters were predicted with the fungal version antiSMASH4.0 program. More than 33 types I polyketide-related biosynthetic gene clusters were distributed, containing PKS and PKS-NRPS (polyketide-nonribosomal peptides) hybrid gene clusters. Another 8 NRPS biosynthetic gene clusters were distributed among the genome of ES026. The prenyltransferases, probably involved in aromatic prenyl-compounds and terpenoid biosynthesis, were analyzed using bioinformatics tools like MEGA. CONCLUSION: We predicted a new possible biosynthetic pathway for the HupA from the pipecolic acid, based on the published HupA biosynthesis proposed pathway, the biosynthesis and pipecolic acid-derived compounds. We hypothesize that a hybrid PKS-NRPS mega-enzyme was probably involved in the biosynthesis of HupA with the pipecolic acid, the building block of rapamycin, as a HupA precursor. The rapamycin is produced from a polyketide biosynthesis pathway, and the domain incorporating the pipecolic acid is studied.
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Colletotrichum , Policetídeos , Colletotrichum/genética , Sequência de Bases , Família Multigênica , Policetídeos/metabolismo , SirolimoRESUMO
For the issue of low accuracy and poor real-time performance of insulator and defect detection by an unmanned aerial vehicle (UAV) in the process of power inspection, an insulator detection model MobileNet_CenterNet was proposed in this study. First, the lightweight network MobileNet V1 was used to replace the feature extraction network Resnet-50 of the original model, aiming to ensure the detection accuracy of the model while speeding up its detection speed. Second, a spatial and channel attention mechanism convolutional block attention module (CBAM) was introduced in CenterNet, aiming to improve the prediction accuracy of small target insulator position information. Then, three transposed convolution modules were added for upsampling, aiming to better restore the semantic information and position information of the image. Finally, the insulator dataset (ID) constructed by ourselves and the public dataset (CPLID) were used for model training and validation, aiming to improve the generalization ability of the model. The experimental results showed that compared with the CenterNet model, MobileNet_CenterNet improved the detection accuracy by 12.2%, the inference speed by 1.1 f/s for FPS-CPU and 4.9 f/s for FPS-GPU, and the model size was reduced by 37 MB. Compared with other models, our proposed model improved both detection accuracy and inference speed, indicating that the MobileNet_CenterNet model had better real-time performance and robustness.
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Actinobacteria are well-known Gram-positive bacteria that produce approximately two-thirds of microbial bioactive natural products (NPs) through secondary metabolism. Usually, genes involved in the biosynthesis of NPs in actinobacteria are clustered, and their expression is regulated by an elaborate and stringent regulatory network formed by diverse regulators. These regulators can be classified into more than 50 superfamilies/families according to conserved amino acid sequences and biological functions. Among them, LuxR family regulators, which are widely distributed in microorganisms and feature an HTH_LUXR domain (PFAM00196, SMART00421), play key roles in quorum sensing (QS), bioluminescence, virulence and secondary metabolism. In this mini-review, we focus on their roles in regulating NP production in actinobacteria. First, the domain architecture and classification of LuxR proteins are summarized on the basis of their size and biological function diversity. Second, the landscape of the roles and action mechanism of LuxR regulators involved in NP production in actinobacteria is presented in detail. Finally, the application of LuxR is described from two perspectives: enhancement of NP production and discovery of novel NPs by engineering LuxR. This mini-review will help us comprehensively understand the role of LuxR in actinobacteria and promote the future application of LuxR family regulators in synthetic biology.
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Actinobacteria , Humanos , Actinobacteria/genética , Actinobacteria/metabolismo , Transativadores/genética , Transativadores/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Percepção de Quorum/genética , Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismoRESUMO
Milbemycins, a group of 16-membered macrocylic lactones with excellent acaricidal, insecticidal and anthelmintic activities, can be produced by several Streptomyces species. For the reason that they have low toxicity in mammals, milbemycins and their derivatives are widely used in agricultural, medical and veterinary industries. Streptomyces bingchenggensis, one of milbemycin-producing strains, has been sequenced and intensively investigated in the past decades. In this mini-review, we comprehensively revisit the progress that has been made in research efforts to elucidate the biosynthetic pathways and regulatory networks for the cellular production of milbemycins. The advances in the development of production strains for milbemycin and its derivatives are discussed along the strain-generation technical approaches of random mutagenesis, metabolic engineering and combinatorial biosynthesis. The research progress made so far indicates that strain improvement and generation of novel milbemycin derivatives will greatly benefit from future development of enabling technologies and deeper understanding of the fundamentals of biosynthesis of milbemycin and the regulation of its production in S. bingchenggensis. This mini-review also proposes that the overproduction of milbemycins could be greatly enhanced by genome minimization, systematical metabolic engineering and synthetic biology approaches in the future.
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Macrolídeos , Streptomyces , Animais , Vias Biossintéticas/genética , Streptomyces/genéticaRESUMO
Streptomyces is famous for its capability to produce the most abundant antibiotics in all kingdoms. All Streptomyces antibiotics are natural products, whose biosynthesis from the so-called gene clusters are elaborately regulated by pyramidal transcriptional regulatory cascades. In the past decades, scientists have striven to unveil the regulatory mechanisms involved in antibiotic production in Streptomyces. Here we mainly focus on three aspects of the regulation on antibiotic production. 1. The onset of antibiotic production triggered by hormones and their coupled receptors as regulators; 2. The cascades of global and pathway-specific regulators governing antibiotic production; 3. The feedback regulation of antibiotics and/or intermediates on the gene cluster expression for their coordinated production. This review will summarize how the antibiotic production is stringently regulated in Streptomyces based on the signaling, and lay a theoretical foundation for improvement of antibiotic production and potentially drug discovery.
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Antibacterianos/metabolismo , Redes Reguladoras de Genes , Streptomyces/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica , Hormônios/metabolismo , Família Multigênica , Streptomyces/genéticaRESUMO
Saccharopolyspora spinosa produces tetra-cyclic macrolide spinosyns, a group of highly efficient pesticidal agents. However, this species lacks efficient vectors for genetic manipulation. In this study, the circular plasmid pCM32 was newly isolated from Saccharopolyspora endophytica YIM 61095. The complete nucleotide sequence of pCM32 consists of 14,611 bp and is predicted to encode 17 open reading frames (ORFs). Interestingly, a putative int gene in pCM32 was predicted by homologous alignment to encode an integrase belonging to the tyrosine family of integrases/recombinases. Plasmid pCM238 containing this int locus derived from pCM32 could be transferred by conjugation from Escherichia coli into Sa. spinosa at a high frequency. Integration of pCM238 in the host chromosome was demonstrated as site-specific recombination (at the tRNA (Ser) gene) via a 56-bp core sequence within the attP/attB sites. Plasmid pCM265, a shuttle vector containing the int and attP sequences of pCM32, was constructed to introduce foreign genes into Sa. spinosa. The production of spinosad approximately doubled in Sa. spinosa NRRL18395 after introducing pCM265-derived plasmids carrying the genes for phosphofructokinase (PFK) or anthranilate synthase. These results indicate that plasmid pCM32 is an actinomycete integrative and conjugative element (AICE) and that its derived integrative vectors are useful for efficiently introducing foreign DNA into Sa. spinosa.
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Macrolídeos/metabolismo , Engenharia Metabólica/métodos , Praguicidas/metabolismo , Plasmídeos , Recombinação Genética , Saccharopolyspora/genética , Saccharopolyspora/metabolismo , Sítios de Ligação Microbiológicos , DNA Bacteriano/química , DNA Bacteriano/genética , Escherichia coli/genética , Expressão Gênica , Vetores Genéticos , Fases de Leitura Aberta , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Análise de Sequência de DNA , Transformação BacterianaRESUMO
Genome mining led to the discovery of a novel aminocoumarin gene cluster in the rare actinomycete Catenulispora acidiphila DSM 44928. Sequence analysis revealed the presence of genes putatively involved in export/resistance, regulation, and biosynthesis of the aminocoumarin moiety and its halogenation, as well as several genes with so far unknown function. Two new aminocoumarins, cacibiocin A and B, were identified in the culture broth of C. acidiphila. Heterologous expression of the putative gene cluster in Streptomyces coelicolor M1152 confirmed that this cluster is responsible for cacibiocin biosynthesis. Furthermore, total production levels of cacibiocins could be increased by heterologous expression and screening of different culture media from an initial yield of 4.9 mg L(-1) in C. acidiphila to 60 mg L(-1) in S. coelicolor M1152. By HR-MS and NMR analysis, cacibiocin A was found to contain a 3-amino-4,7-dihydroxycoumarin moiety linked by an amide bond to a pyrrole-2,5-dicarboxylic acid. The latter structural motif has not been identified previously in any natural compound. Additionally, cacibiocin B contains two chlorine atoms at positions 6' and 8' of the aminocoumarin moiety.
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Actinomycetales/química , Aminocumarinas/química , Antibacterianos/química , Pirróis/química , Actinomycetales/genética , Actinomycetales/metabolismo , Aminocumarinas/metabolismo , Aminocumarinas/farmacologia , Antibacterianos/biossíntese , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Produtos Biológicos/química , Produtos Biológicos/metabolismo , Produtos Biológicos/farmacologia , Genoma Bacteriano , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Conformação Molecular , Família Multigênica , Pirróis/metabolismo , Pirróis/farmacologia , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Streptomyces/metabolismoRESUMO
Streptomyces species are best known for their ability to produce abundant secondary metabolites with versatile bioactivities and industrial importance. These metabolites are usually biosynthesized through metabolic pathways encoded by cluster-situated genes. These genes are also known as biosynthetic gene clusters (BGCs) of secondary metabolites. The expression of BGCs is intricately controlled by pyramidal transcriptional regulatory cascades, which include various regulators. Streptomyces antibiotic regulatory proteins (SARPs), a genus-specific family of regulators, are widely distributed and play important roles in regulating the biosynthesis of secondary metabolites in Streptomyces. Over the past decade, the biological functions of SARPs have been extensively investigated. Here, we summarized the recent advances in characterizing the roles of SARPs involved in Streptomyces secondary metabolism from the following three aspects. First, the classification and domain organization of SARPs were summarized according to their size variation. Second, we presented a detailed description of the regulatory mechanisms and modes of action of SARPs involved in secondary metabolism. Finally, the biotechnological application of SARPs was illustrated by improving the production of target secondary metabolites and discovering novel bioactive natural products. This review will help researchers to comprehensively understand the roles of SARPs in secondary metabolite biosynthesis in Streptomyces, which will contribute to building a solid foundation for their future application in synthetic biology.
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Generative adversarial network (GAN) has achieved remarkable success in generating high-quality synthetic data by learning the underlying distributions of target data. Recent efforts have been devoted to utilizing optimal transport (OT) to tackle the gradient vanishing and instability issues in GAN. They use the Wasserstein distance as a metric to measure the discrepancy between the generator distribution and the real data distribution. However, most optimal transport GANs define loss functions in Euclidean space, which limits their capability in handling high-order statistics that are of much interest in a variety of practical applications. In this article, we propose a computational framework to alleviate this issue from both theoretical and practical perspectives. Particularly, we generalize the optimal transport-based GAN from Euclidean space to the reproducing kernel Hilbert space (RKHS) and propose Hilbert Optimal Transport GAN (HOT-GAN). First, we design HOT-GAN with a Hilbert embedding that allows the discriminator to tackle more informative and high-order statistics in RKHS. Second, we prove that HOT-GAN has a closed-form kernel reformulation in RKHS that can achieve a tractable objective under the GAN framework. Third, HOT-GAN's objective enjoys the theoretical guarantee of differentiability with respect to generator parameters, which is beneficial to learn powerful generators via adversarial kernel learning. Extensive experiments are conducted, showing that our proposed HOT-GAN consistently outperforms the representative GAN works.
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Soil salinization is one of the major soil degradation threats worldwide, and parameters related to soil quality and ecosystem multifunctionality (EMF) are crucial for evaluating the success of reclamation efforts in saline-sodic wasteland (WL). Microbial metabolic limitation is also one of the main factors that influences EMF in agricultural cropping systems. A ten-year localization experiment was conducted to reveal the key predictors of soil quality index (SQI) values, microbial metabolic characteristics, and EMF in different farmland cropping systems. A random forest model showed that the ß-glucosidase (BG), cellobiosidase (CBH) and saturated hydraulic conductivity (SHC) of the SQI factors were the main driving forces of soil EMF. Compared to monoculture models, such as paddy field (PF) or upland field (UF), the converted paddy field to upland field (CF) cropping system was most effective at improving EMF in reclaimed saline-sodic WL, increasing this metric by 275.35 %. CF integrates practices from both PF and UF planting systems, improved soil quality and relieves microbial metabolic limitation. Specifically, both CF and PF significantly reduced soil pH (by 16-23 %) and sodium adsorption ration (SAR) (by 65-83 %) and significantly reduced the abundance of large macroaggregates. Moreover, CF significantly improved soil saturated hydraulic conductivity relative to PF and UF (p < 0.05), indicating an improvement in soil physical properties. Overall, although reclamation improved SQI compared to WL (0.25), the EMF of CF (0.56) was significantly higher than that of other treatments (p < 0.05). Thus, while increasing SQI can improve soil EMF, it was not as effective alone as it was when combined with more comprehensive efforts that focus on improving various soil properties and alleviating microbial metabolic limitations. Therefore, our results suggested that future saline-sodic wasteland reclamation efforts should avoid monoculture systems to enhance soil EMF.
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Ecossistema , Solo , Solo/química , Sódio/química , AdsorçãoRESUMO
Avermectins (AVMs), a family of 16-membered macrocyclic macrolides produced by Streptomyces avermitilis, have been the most successful microbial natural antiparasitic agents in recent decades. Doramectin, an AVM derivative produced by S. avermitilis bkd- mutants through cyclohexanecarboxylic acid (CHC) feeding, was commercialized as a veterinary antiparasitic drug by Pfizer Inc. Our previous results show that the production of avermectin and actinorhodin was affected by several other polyketide biosynthetic gene clusters in S. avermitilis and Streptomyces coelicolor, respectively. Thus, here, we propose a rational strategy to improve doramectin production via the termination of competing polyketide biosynthetic pathways combined with the overexpression of CoA ligase, providing precursors for polyketide biosynthesis. fadD17, an annotated putative cyclohex-1-ene-1-carboxylate:CoA ligase-encoding gene, was proven to be involved in the biosynthesis of doramectin. By sequentially removing three PKS (polyketide synthase) gene clusters and overexpressing FadD17 in the strain DM203, the resulting strain DM223 produced approximately 723 mg/L of doramectin in flasks, which was approximately 260% that of the original strain DM203 (approximately 280 mg/L). To summarize, our work demonstrates a novel viable approach to engineer doramectin overproducers, which might contribute to the reduction in the cost of this valuable compound in the future.
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The genus of Nocardiopsis is a new source of antibiotics, chemicals, and enzymes. Here we reported the development of a vector and host system in moderately halophilic Nocardiopsis via an oriT-mediated conjugation. By screening about 80 Nocardiopsis strains, 6 of them harbored 8 plasmids (18-80 kb). The complete nucleotide sequence of pSQ10 consisted of 18,219 bp, with 71.9% G + C content, encoding 17 open reading frames, 5 of them resembled those of Streptomyces plasmids. A rep locus (iteron within the gene) was identified for replication in Nocardiopsis sp. YIM 90083, and rep protein bound to its iteron sequence. This system may be useful for gene cloning and expression in Nocardiopsis.