1,646 results match your criteria Acs Synthetic Biology[Journal]


Reprogramming acetogenic bacteria with CRISPR-targeted base editing via deamination.

ACS Synth Biol 2020 Jul 1. Epub 2020 Jul 1.

Acetogenic bacteria are rising in popularity as chassis microbes in biotechnology due to their capability of converting inorganic one-carbon (C1) gases to organic chemicals. To fully uncover the potential of acetogenic bacteria, synthetic-biology tools are imperative to either engineer designed functions or to interrogate the physiology. Here, we report a genome-editing tool at a one-nucleotide resolution, namely base editing, for acetogenic bacteria based on CRISPR-targeted deamination. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00226DOI Listing

Orthogonal Blue and Red Light Controlled Cell-Cell Adhesions enable Sorting-out in Multicellular Structures.

ACS Synth Biol 2020 Jul 1. Epub 2020 Jul 1.

The self-assembly of different cell types into multicellular structures and their organization into spatiotemporally controlled patterns are both challenging and extremely powerful to understand how cells function within tissues and for bottom-up tissue engineering. Here, we not only independently control the self-assembly of two cell types into multicellular architectures with blue and red light, but also achieve their self-sorting into distinct assemblies. This required developing two cell types that form selective and homophilic cell-cell interactions either under blue or red light using photoswitchable proteins as artificial adhesion molecules. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00150DOI Listing

Identification of the biosynthetic gene cluster for the anti-MRSA lysocins through gene cluster activation using strong promoters of housekeeping genes and production of new analogs in sp. 3655.

ACS Synth Biol 2020 Jul 1. Epub 2020 Jul 1.

The Gram-negative gliding bacteria represent a new and rich source for bioactive natural products. In an effort to discover new antibiotics, we found a cryptic biosynthetic gene cluster (BGC) in sp. 3655 that shared a high similarity with the putative lysocin BGC identified previously from sp. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00067DOI Listing

Reassembly of the biosynthetic gene cluster enables high epothilone yield in engineered Schlegelella brevitalea.

ACS Synth Biol 2020 Jun 30. Epub 2020 Jun 30.

Epothilones, as a new class of microtubule-stabilizing anticancer drugs, exhibit strong bioactivity against taxane-resistant cells and clinically show active for the treatment of advanced breast cancer. Additionally, they also imply great potential values for central nervous system injury and Alzheimer's disease. However, due to the long fermentation period of the original producer and challenges of genetic engineering of nonribosomal peptide/polyketide (NRP/PK) megasynthase genes, the application of epothilones is severely limited. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00100DOI Listing

Bringing light into cell-free expression.

ACS Synth Biol 2020 Jun 30. Epub 2020 Jun 30.

Cell-free system, as part of the synthetic biology field, has become a critical platform in the biological studies. However, there is a lack of research into developing a switch for a dynamical control of the transcriptional and translational process. The optogenetic tool has been widely proven as an ideal control switch for protein synthesis due to its non-toxicity and excellent time-space conversion. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00211DOI Listing

Yeast Intracellular Staining (yICS): Enabling High-throughput, Quantitative Detection of Intracellular Proteins via Flow Cytometry for Pathway Engineering.

ACS Synth Biol 2020 Jun 30. Epub 2020 Jun 30.

The complexities of pathway engineering necessitate screening libraries to discover phenotypes of interest. However, this approach is challenging when desirable phenotypes cannot be directly linked to growth advantages or fluorescence. In these cases, the ability to rapidly quantify intracellular proteins in the pathway of interest is critical to expedite the clonal selection process. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00199DOI Listing

Optimal Ratio of Carbon Flux between Glycolysis and the Pentose Phosphate Pathway for Amino Acid Accumulation in .

ACS Synth Biol 2020 Jun 30. Epub 2020 Jun 30.

Graduate School of Natural Science and Technology, Kanazawa University, Kakuma-machi, Kanazawa, Ishikawa 920-1192, Japan.

Glucose is metabolized through central metabolic pathways such as glycolysis and the pentose phosphate pathway (PPP) to synthesize downstream metabolites including amino acids. However, how the split ratio of carbon flux between glycolysis and PPP specifically affects the formation of downstream metabolites remains largely unclear. Here, we conducted a comprehensive metabolomic analysis to investigate the effect of the split ratio between glycolysis and the PPP on the intracellular concentration of amino acids and their derivatives in . Read More

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http://dx.doi.org/10.1021/acssynbio.0c00181DOI Listing

Domain-specific programming languages for computational nucleic acid systems.

ACS Synth Biol 2020 Jun 26. Epub 2020 Jun 26.

The construction of models of system behavior is of great importance throughout science and engineering. In bioengineering and bionanotechnology, these often take the form of dynamic models that specify the evolution of different species over time. In order to ensure that scientific observations and conclusions are consistent, and that systems can be reliably engineered on the basis of model predictions, it is important that models of biomolecular systems can be constructed in a reliable, principled, and efficient manner. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00050DOI Listing

Distributed implementation of Boolean functions by transcriptional synthetic circuits.

ACS Synth Biol 2020 Jun 26. Epub 2020 Jun 26.

Starting in the early 2000s, sophisticated technology has been developed for the rational construction of synthetic genetic networks that implement specified logical functionalities. Despite impressive progress, however, the scaling necessary in order to achieve greater computational power has been hampered by many constraints, including repressor toxicity and the lack of large sets of mutually-orthogonal repressors. As a consequence, a typical circuit contains no more than roughly seven repressor-based gates per cell. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00228DOI Listing

Engineering Carboxylic Acid Reductase (CAR) through A Whole-Cell Growth-Coupled NADPH Recycling Strategy.

ACS Synth Biol 2020 Jun 26. Epub 2020 Jun 26.

Rapid evolution of enzyme activities is often hindered by the lack of efficient and affordable methods to identify beneficial mutants. We report the development of a new growth-coupled selection method for evolving NADPH-consuming enzymes based on the recycling of this redox cofactor. The method relies on a genetically modified Escherichia coli strain, which overaccumulates NADPH. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00290DOI Listing

Robotic DNA Nanostructures.

ACS Synth Biol 2020 Jun 26. Epub 2020 Jun 26.

Over the last decade, DNA nanotechnology has spawned a broad variety of functional nanostructures tailored toward the enabled state at which applications are coming increasingly in view. One of the branches of these applications is in synthetic biology, where the intrinsic programmability of the DNA nanostructures may pave the way for smart task-specific molecular robotics. In brief, the synthesis of the user-defined artificial DNA nano-objects is based on employing DNA molecules with custom lengths and sequences as building materials that predictably assemble together by obeying Watson-Crick base pairing rules. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00235DOI Listing

Metabolic engineering of Saccharomyces cerevisiae for rosmarinic acid production.

ACS Synth Biol 2020 Jun 26. Epub 2020 Jun 26.

Rosmarinic acid is a hydroxycinnamic acid ester commonly found in the Boraginaceae and Lamiaceae plant families. It exhibits various biological activities, including antioxidant, anti-inflammatory, antibacterial, anti-allergic, and antiviral properties. Rosmarinic acid is used as a food and cosmetic ingredient and several pharmaceutical applications have been suggested as well. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00048DOI Listing

Processing DNA-Based Molecular Signals into Graphical Displays.

ACS Synth Biol 2020 Jun 25. Epub 2020 Jun 25.

The RNA Institute, University at Albany, State University of New York, Albany, New York 12222, United States.

DNA is now well-established as a nanoscale building material with applications in fields such as biosensing and molecular computation. Molecular processes such as logic gates, nucleic acid circuits, and multiplexed detection have used different readout strategies to measure the output signal. In biosensing, this output can be the diagnosis of a disease biomarker, whereas in molecular computation, the output can be the result of a mathematical operation carried out using DNA. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00246DOI Listing

Expanding Toolbox for Genes Expression of Yarrowia lipolytica to Include Novel Inducible, Repressible and Hybrid Promoters.

ACS Synth Biol 2020 Jun 25. Epub 2020 Jun 25.

Promoters are critical tools to precisely control genes expression for both synthetic biology and metabolic engineering. Although Yarrowia lipolytica has demonstrated with many industrially relevant advantages, promoter discovery efforts on this non-conventional yeast are limited due to the challenge in finding suitable inducible and repressible promoters. Six copper-inducible promoters and five repressible promoters were isolated in this work. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00243DOI Listing

Site-directed mutagenesis of large biosynthetic gene clusters via oligonucleotide recombineering and CRISPR/Cas9 targeting.

ACS Synth Biol 2020 Jun 25. Epub 2020 Jun 25.

Genetic engineering of natural product biosynthetic gene clusters represents an attractive approach to access new and complex bioactive small molecules. However, due to the large number and size of some genes involved in specialized metabolism, notably those encoding modular polyketide synthase and nonribosomal peptide synthetase megaproteins, it remains difficult to introduce precise genetic mutations to probe domain activity or alter chemical product formation. Here, we report the development and validation of a robust method combining oligonucleotide recombineering and CRISPR/Cas9 targeting for rapid site-directed mutagenesis of cloned pathways, which can be directly transferred to a heterologous host for expression. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00265DOI Listing

Purification of an Intact Human Protein Overexpressed from Its Endogenous Locus Direct Genome Engineering.

ACS Synth Biol 2020 Jul 2. Epub 2020 Jul 2.

Department of Chemistry, Hanyang University, Seoul 04763, South Korea.

The overproduction and purification of human proteins is a requisite of both basic and medical research. Although many recombinant human proteins have been purified, current protein production methods have several limitations; recombinant proteins are frequently truncated, fail to fold properly, and/or lack appropriate post-translational modifications. In addition, such methods require subcloning of the target gene into relevant plasmids, which can be difficult for long proteins with repeated domains. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00090DOI Listing

Identification of a strong quorum sensing- and thermo-regulated promoter for the biosynthesis of a new metabolite pesticide phenazine-1-carboxamide in Pseudomonas strain PA1201.

ACS Synth Biol 2020 Jun 25. Epub 2020 Jun 25.

Phenazine-1-carboxamide (PCN) produced by multifarious Pseudomonas strains represents a promising candidate as a new metabolite pesticide due to its broad-spectrum antifungal activity and capacity to induce systemic resistance in plants. The rice rhizosphere Pseudomonas strain PA1201 contains two reiterated gene clusters, phz1 and phz2, for phenazine-1-carboxylic acid (PCA) biosynthesis; PCA is further converted into PCN by this strain using a functional phzH-encoding glutamine aminotransferase. However, PCN levels in PA1201 constitute approximately one-fifth of PCA levels and the optimal temperature for PCN synthesis is 28C. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00161DOI Listing

Exploring Catalysis Specificity of Phytoene Dehydrogenase CrtI in Carotenoid Synthesis.

ACS Synth Biol 2020 Jun 24. Epub 2020 Jun 24.

Carotenoids, a variety of natural products, have significant pharmaceutical and commercial potential. Phytoene dehydrogenase (CrtI) is the rate-limit enzyme for carotenoid synthesis, whose catalysis specificity results in various carotenoids. However, the structural characteristics of CrtI for controlling the catalysis specificity on dehydrogenation steps are still unclear, which limited the development of CrtI function. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00128DOI Listing

Synthetic Control of Signal Flow Within a Bacterial Multi-Kinase Network.

ACS Synth Biol 2020 Jun 19. Epub 2020 Jun 19.

The signal processing capabilities of bacterial signaling networks offer immense potential for advanced phospho-signaling systems for synthetic biology. Emerging models suggest that complex development may require interconnections between what were once thought to be isolated signaling arrays. For example, Caulobacter crescentus achieves the feat of asymmetric division by utilizing a novel pseudokinase DivL, which senses the output of one signaling pathway to modulate a second pathway. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00043DOI Listing

Preparation of a Millimeter-Sized Supergiant Liposome That Allows for Efficient, Eukaryotic Cell-Free Translation in the Interior by Spontaneous Emulsion Transfer.

ACS Synth Biol 2020 Jun 26. Epub 2020 Jun 26.

Proteo-Science Center, Ehime University, 3 Bunkyo-cho, Matsuyama, Ehime 790-8577, Japan.

We sought to prepare millimeter-sized supergiant unilamellar vesicles (SGUVs) by spontaneous emulsion transfer for efficient, eukaryotic cell-free translation in the interior. Although the conventional protocols require that a considerably high concentration of sucrose be encapsulated into the SGUVs for their efficient formation, such high amounts of sucrose severely inhibited cell-free translation based on wheat germ extract (WGE). We thus optimized the preparation conditions to permit SGUV formation at a much lower concentration of sucrose that has almost no effect on WGE translation. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00173DOI Listing

Synthesis Success Calculator: Predicting the Rapid Synthesis of DNA Fragments with Machine Learning.

ACS Synth Biol 2020 Jun 30. Epub 2020 Jun 30.

The synthesis and assembly of long DNA fragments has greatly accelerated synthetic biology and biotechnology research. However, long turnaround times or synthesis failures create unpredictable bottlenecks in the design-build-test-learn cycle. We developed a machine learning model, called the Synthesis Success Calculator, to predict whether a long DNA fragment can be readily synthesized with a short turnaround time. Read More

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http://dx.doi.org/10.1021/acssynbio.9b00460DOI Listing

Multiplex evolution of antibody fragments utilizing a yeast surface display platform.

ACS Synth Biol 2020 Jun 18. Epub 2020 Jun 18.

Advances in high-throughput synthetic biology technologies based on the CRISPR/Cas9 system have enabled a comprehensive assessment of mutations conferring desired phenotypes, as well as a better understanding of genotype-phenotype correlations in protein engineering. Engineering antibodies to enhance properties such as binding affinity and stability plays an essential role in therapeutic applications. Here we report a method, multiplex navigation of antibody structure (MINAS), that combines a CRISPR/Cas9-based trackable editing method and fluorescent-activated cell sorting (FACS) of yeast-displayed libraries. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00159DOI Listing

Directed evolution of ornithine cyclodeaminase using an EvolvR-based growth-coupling strategy for efficient biosynthesis of L-proline.

ACS Synth Biol 2020 Jun 18. Epub 2020 Jun 18.

L-proline takes a significant role in pharmaceutical and chemical industry as well as graziery. Typical biosynthesis of L-proline is from L-glutamate, involving three enzymes reaction as well as a spontaneous cyclisation. Alternatively, L-proline can be also synthesized in L-ornithine and/or L-arginine producing strains by an ornithine aminotransferase (OCD). Read More

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http://dx.doi.org/10.1021/acssynbio.0c00198DOI Listing

CRISPR-dCas9 Mediated Cytosine Deaminase Base Editing in .

ACS Synth Biol 2020 Jul 1. Epub 2020 Jul 1.

College of Life Sciences, University of Chinese Academy of Sciences, Beijing 100049, China.

Base editing technology based on clustered regularly interspaced short palindromic repeats/associated protein 9 (CRISPR/Cas9) is a recent addition to the family of CRISPR technologies. Compared with the traditional CRISPR/Cas9 technology, it does not rely on DNA double strand break and homologous recombination, and can realize gene inactivation and point mutation more quickly and simply. Herein, we first developed a base editing method for genome editing in utilizing CRISPR/dCas9 (a fully nuclease-deficient mutant of Cas9 from ) and activation-induced cytidine deaminase (AID). Read More

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http://dx.doi.org/10.1021/acssynbio.0c00151DOI Listing

A Library of Tunable, Portable, and Inducer-Free Promoters Derived from Cyanobacteria.

ACS Synth Biol 2020 Jul 1. Epub 2020 Jul 1.

Cyanobacteria are emerging as hosts for various biotechnological applications. The ability to engineer these photosynthetic prokaryotes greatly depends on the availability of well-characterized promoters. Inducer-free promoters of a range of activities may be desirable for the eventual large-scale, outdoor cultivations. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00152DOI Listing

Mini PG10 as a Convenient and Effective Production Host for Lantibiotics.

ACS Synth Biol 2020 Jun 30. Epub 2020 Jun 30.

Department of Molecular Genetics, University of Groningen, Groningen, 9747AG, The Netherlands.

Efficient bacterial cell factories are important for the screening and characterization of potent antimicrobial peptides such as lantibiotics. Although lantibiotic production systems have been established in and , the industrial workhorse has been left relatively unexplored as a lantibiotic production host. Therefore, we tested different strains for their ability to produce lantibiotic peptides by using the subtilin modification and transport enzymes derived from the natural subtilin producer ATCC 6633. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00194DOI Listing

Controlled functional zonation of hepatocytes in vitro by engineering of Wnt signalling.

ACS Synth Biol 2020 Jun 18. Epub 2020 Jun 18.

Key liver functions including protein synthesis, carbohydrate metabolism and detoxification are performed by specific populations of hepatocytes which are defined by their relative position within the liver lobules. On a molecular level, the functional heterogeneity with periportal and pericentral phenotypes, so-called metabolic liver zonation, is mainly established by a gradient of canonical Wnt-signalling activity. Since the relevant physiological cues are missing in in vitro liver models, they fail to reflect the functional heterogeneity and thus lack many liver functions. Read More

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http://dx.doi.org/10.1021/acssynbio.9b00435DOI Listing

Rational Design and Self-Assembly of Coiled-Coil Linked SasG Protein Fibrils.

ACS Synth Biol 2020 Jun 22. Epub 2020 Jun 22.

Department of Chemistry, University of Sheffield, Sheffield S3 7HF, United Kingdom.

Protein engineering is an attractive approach for the self-assembly of nanometer-scale architectures for a range of potential nanotechnologies. Using the versatile chemistry provided by protein folding and assembly, coupled with amino acid side-chain functionality, allows for the construction of precise molecular "protein origami" hierarchical patterned structures for a range of nanoapplications such as stand-alone enzymatic pathways and molecular machines. The surface protein SasG is a rigid, rod-like structure shown to have high mechanical strength due to "clamp-like" intradomain features and a stabilizing interface between the G5 and E domains, making it an excellent building block for molecular self-assembly. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00156DOI Listing

Functional integration of two CYP450 genes involved in biosynthesis of tanshinones for improved diterpenoid production by synthetic biology.

ACS Synth Biol 2020 Jun 18. Epub 2020 Jun 18.

Cytochrome P450s (CYPs) are important enzymes in the secondary metabolism of plants and have been recognized as key players in bioengineering and synthetic biology. Previously reported CYP76AH1 and CYP76AH3, having greater than 80% sequence homology that played a continuous catalytic role in the biosynthesis of tanshinones in . Homology modeling indicates that four sites might be responsible for differences in catalytic activity between the two enzymes. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00136DOI Listing

Multicopy chromosomal integration using CRISPR-associated transposases.

ACS Synth Biol 2020 Jun 17. Epub 2020 Jun 17.

Controlling the copy number of gene expression cassettes is an important strategy to engineer bacterial cells into high-efficient biocatalysts. Current strategies mostly use plasmid vectors, but multicopy plasmids are often genetically unstable, and their copy numbers cannot be precisely controlled. The integration of expression cassettes into a bacterial chromosome has advantages, but iterative integration is laborious, and it is challenging to obtain a library with varied gene doses for phenotype characterization. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00073DOI Listing

Robust Heterochiral Strand Displacement Using Leakless Translators.

ACS Synth Biol 2020 Jul 1. Epub 2020 Jul 1.

Center for Biomedical Engineering, University of New Mexico, Albuquerque, New Mexico 87131, United States.

Molecular computing offers a powerful framework for biosensing and signal processing at the nanoscale. However, for applications, the use of conventional DNA components can lead to false positive signals being generated due to degradation of circuit components by nuclease enzymes. Here, we use hybrid chiral molecules, consisting of both l- and d-nucleic acid domains, to implement leakless signal translators that enable d-nucleic acid signals to be detected by hybridization and then translated into a robust l-DNA signal for further analysis. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00131DOI Listing

Endogenous p21-Dependent Transgene Control for CHO Cell Engineering.

ACS Synth Biol 2020 Jun 15. Epub 2020 Jun 15.

Department of Molecular Science and Technology, Ajou University, Suwon 16499, Republic of Korea.

Numerous engineering efforts have been made in Chinese hamster ovary (CHO) cells for high level production of therapeutic proteins. However, the dynamic regulation of transgene expression is limited in current systems. Here, we investigated the effective regulation of transgene expression in CHO cells targeted integration-based endogenous gene tagging with engineering target genes. Read More

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http://dx.doi.org/10.1021/acssynbio.9b00526DOI Listing
June 2020
3.951 Impact Factor

A dual-plasmid CRISPR/Cas system for mycotoxin elimination in polykaryotic industrial fungi.

ACS Synth Biol 2020 Jun 12. Epub 2020 Jun 12.

Mycotoxin contamination causes disease and death in both humans and animals. Monascus Red, produced by Monascus purpureus, is used as a food colorant. However, its application is limited by contamination of the nephrotoxin citrinin, which is also produced by the fungus. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00178DOI Listing

Artificial Protein-Responsive Riboswitches Upregulate Non-AUG Translation Initiation in Yeast.

ACS Synth Biol 2020 Jun 23. Epub 2020 Jun 23.

Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, 5-1-5 Kashiwanoha, Kashiwa-shi, Chiba 277-8562, Japan.

Artificial control of gene expression is one of the core technologies for engineering biological systems. Riboswitches are cis-acting elements on mRNA that regulate gene expression in a ligand-dependent manner often seen in prokaryotes, but rarely in eukaryotes. Because of the poor variety of such elements available in eukaryotic systems, the number of artificially engineered eukaryotic riboswitches, especially of the upregulation type, is still limited. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00206DOI Listing

Programmed Allelic Mutagenesis of a DNA Polymerase with Single Amino Acid Resolution.

ACS Synth Biol 2020 Jun 30. Epub 2020 Jun 30.

Department of Pharmaceutical Sciences, University of California, Irvine, California 92697-3958, United States.

Most DNA polymerase libraries sample unknown portions of mutational space and are constrained by the limitations of random mutagenesis. Here we describe a programmed allelic mutagenesis (PAM) strategy to comprehensively evaluate all possible single-point mutations in the entire catalytic domain of a replicative DNA polymerase. By applying the PAM strategy with ultrafast high-throughput screening, we show how DNA polymerases can be mapped for allelic mutations that exhibit enhanced activity for unnatural nucleic acid substrates. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00236DOI Listing

Synthetic Glycobiology: Parts, Systems, and Applications.

ACS Synth Biol 2020 Jun 30. Epub 2020 Jun 30.

Department of Chemical and Biological Engineering, Northwestern University, 2145 Sheridan Road, Tech E136, Evanston, Illinois 60208, United States.

Protein glycosylation, the attachment of sugars to amino acid side chains, can endow proteins with a wide variety of properties of great interest to the engineering biology community. However, natural glycosylation systems are limited in the diversity of glycoproteins they can synthesize, the scale at which they can be harnessed for biotechnology, and the homogeneity of glycoprotein structures they can produce. Here we provide an overview of the emerging field of synthetic glycobiology, the application of synthetic biology tools and design principles to better understand and engineer glycosylation. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00210DOI Listing

CRISPR-Mediated Activation of Biosynthetic Gene Clusters for Bioactive Molecule Discovery in Filamentous Fungi.

ACS Synth Biol 2020 Jun 26. Epub 2020 Jun 26.

School of Molecular Sciences, University of Western Australia, Perth, WA 6009, Australia.

Accessing the full biosynthetic potential encoded in the genomes of fungi is limited by the low expression of most biosynthetic gene clusters (BGCs) under common laboratory culture conditions. CRISPR-mediated transcriptional activation (CRISPRa) of fungal BGCs could accelerate genomics-driven bioactive secondary metabolite discovery. In this work, we established the first CRISPRa system for filamentous fungi. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00197DOI Listing

Assessment of Robustness to Temperature in a Negative Feedback Loop and a Feedforward Loop.

ACS Synth Biol 2020 Jun 25. Epub 2020 Jun 25.

Department of Electrical Engineering, Indian Institute of Technology Delhi, Hauz Khas, New Delhi, 110016, India.

Robustness to temperature variation is an important specification in biomolecular circuit design. While the cancellation of parametric temperature dependencies has been shown to improve the temperature robustness of the period in a synthetic oscillator design, the performance of other biomolecular circuit designs in different temperature conditions is relatively unclear. Using a combination of experimental measurements and mathematical models, we assessed the temperature robustness of two biomolecular circuit motifs-a negative feedback loop and a feedforward loop. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00023DOI Listing

Rational Design of Flavonoid Production Routes Using Combinatorial and Precursor-Directed Biosynthesis.

ACS Synth Biol 2020 Jun 27. Epub 2020 Jun 27.

Leibniz Research Cluster Group "Biobricks of Microbial Natural Product Syntheses", Leibniz Institute for Natural Product Research and Infection Biology - Hans Knöll Institute, Jena 07745, Germany.

Combinatorial biosynthesis has great potential for designing synthetic circuits and amplifying the production of new active compounds. Studies on multienzyme cascades are extremely useful for improving our knowledge on enzymatic catalysis. In particular, the elucidation of enzyme substrate promiscuity can be potentially used for bioretrosynthetic approaches, leading to the design of alternative and more convenient routes to produce relevant molecules. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00172DOI Listing

From Electronic Sequence to Purified Protein Using Automated Gene Synthesis and Transcription/Translation.

ACS Synth Biol 2020 Jun 25. Epub 2020 Jun 25.

Thermo Fisher Scientific, GENEART GmbH, Im Gewerbepark B35, 93059 Regensburg, Germany.

gene synthesis is the state-of-the-art method used to obtain genetic material adapted to the requirements of the host organism and a cornerstone for modern synthetic biology. Yet, little progress has been made regarding downstream processes of protein production from synthetic genetic material. The production of recombinant proteins traditionally requires extensive preparatory work including gene amplification, cloning, sequencing, transformation or transfection of the expression host, cultivation of living cells, and purification of the overexpressed protein. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00060DOI Listing

Genome Editing Based on Oligo Recombineering and Cas9-Mediated Counterselection.

ACS Synth Biol 2020 Jun 22. Epub 2020 Jun 22.

Centre for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology, Dr. Aiguader 88, Barcelona 08003, Spain.

Mycoplasma species share a set of features, such as lack of a cell wall, streamlined genomes, simplified metabolism, and the use of a deviant genetic code, that make them attractive approximations of what a chassis strain should ideally be. Among them, arises as a candidate for synthetic biology projects, as it is one of the most deeply characterized bacteria. However, the historical paucity of tools for editing Mycoplasma genomes has precluded the establishment of as a suitable chassis strain. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00022DOI Listing

Mapping Protein-Protein Interaction Interface Peptides with Jun-Fos Assisted Phage Display and Deep Sequencing.

ACS Synth Biol 2020 Jun 23. Epub 2020 Jun 23.

Protein-protein interactions govern many cellular processes, and identifying binding interaction sites on proteins can facilitate the discovery of inhibitors to block such interactions. Here we identify peptides from a randomly fragmented plasmid encoding the β-lactamase inhibitory protein (BLIP) and the Lac repressor (LacI) that represent regions of protein-protein interactions. We utilized a Jun-Fos-assisted phage display system that has previously been used to screen cDNA and genomic libraries to identify antibody antigens. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00242DOI Listing

Microbial Synthesis of Human-Hormone Melatonin at Gram Scales.

ACS Synth Biol 2020 Jun 5;9(6):1240-1245. Epub 2020 Jun 5.

Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Kongens Lyngby 2800, Denmark.

Melatonin is a commercially attractive tryptophan-derived hormone. Here we describe a bioprocess for the production of melatonin using to high titers. The first engineered strain produced 0. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00065DOI Listing

Evolving a Thermostable Terminal Deoxynucleotidyl Transferase.

ACS Synth Biol 2020 Jun 23. Epub 2020 Jun 23.

Illumina Singapore Pte, Ltd., 29 Woodlands Industrial Park E1, North Tech Building, Singapore 757716.

Terminal deoxynucleotidyl transferase (TdT) catalyzes template free incorporation of arbitrary nucleotides onto single-stranded DNA. Due to this unique feature, TdT is widely used in biotechnology and clinical applications. One particularly tantalizing use is the synthesis of long DNA molecules by TdT-mediated iterative incorporation of a 3' reversibly blocked nucleotide, followed by deblocking. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00078DOI Listing

Development of Novel Riboswitches for Synthetic Biology in the Green Alga Chlamydomonas.

ACS Synth Biol 2020 Jun 4;9(6):1406-1417. Epub 2020 Jun 4.

Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, U.K.

Riboswitches are RNA regulatory elements that bind specific ligands to control gene expression. Because of their modular composition, where a ligand-sensing aptamer domain is combined with an expression platform, riboswitches offer unique tools for synthetic biology applications. Here we took a mutational approach to determine functionally important nucleotide residues in the thiamine pyrophosphate (TPP) riboswitch in the THI4 gene of the model alga , allowing us to carry out aptamer swap using THIC aptamers from and . Read More

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http://dx.doi.org/10.1021/acssynbio.0c00082DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7309327PMC

Pattern Engineering of Living Bacterial Colonies Using Meniscus-Driven Fluidic Channels.

ACS Synth Biol 2020 Jun 3;9(6):1277-1283. Epub 2020 Jun 3.

School of Life Sciences, University of Warwick, Coventry CV4 7AL, United Kingdom.

Creating adaptive, sustainable, and dynamic biomaterials is a forthcoming mission of synthetic biology. Engineering spatially organized bacterial communities has a potential to develop such bio-metamaterials. However, generating living patterns with precision, robustness, and a low technical barrier remains as a challenge. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00146DOI Listing

Biosystems Design by Machine Learning.

ACS Synth Biol 2020 Jun 29. Epub 2020 Jun 29.

Biosystems such as enzymes, pathways, and whole cells have been increasingly explored for biotechnological applications. However, the intricate connectivity and resulting complexity of biosystems poses a major hurdle in designing biosystems with desirable features. As -omics and other high throughput technologies have been rapidly developed, the promise of applying machine learning (ML) techniques in biosystems design has started to become a reality. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00129DOI Listing

Dynamic Cell Programming with Quorum Sensing-Controlled CRISPRi Circuit.

ACS Synth Biol 2020 Jun 5;9(6):1284-1291. Epub 2020 Jun 5.

Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, Ontario M5S 3E5, Canada.

Synthetic biology is enabling rapid advances in the areas of biomanufacturing and live therapeutics. Dynamic circuits that can be used to regulate cellular resources and microbial community behavior represent a defining focus of synthetic biology, and have attracted tremendous interest. However, the existing dynamic circuits are mostly gene editing-dependent or cell lysis-based, which limits their broad and convenient application, and in some cases, such lysis-based circuits can suffer from genetic instability due to evolution. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00148DOI Listing

Targeted Diversification in the Genome with CRISPR-Guided DNA Polymerase I.

ACS Synth Biol 2020 Jun 16. Epub 2020 Jun 16.

Department of Bioengineering, University of California, Berkeley, California 94720, United States.

New technologies to target nucleotide diversification are promising enabling strategies to perform directed evolution for engineering applications and forward genetics for addressing biological questions. Recently, we reported EvolvR-a system that employs CRISPR-guided Cas9 nickases fused to nick-translating, error-prone DNA polymerases to diversify targeted genomic loci-in . As CRISPR-Cas9 has shown activity across diverse cell types, EvolvR has the potential to be ported into other organisms, including eukaryotes, if nick-translating polymerases can be active across species. Read More

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http://dx.doi.org/10.1021/acssynbio.0c00149DOI Listing