Abstract：
:To spatially control biochemical functions at specific sites within a genome, we have engineered a synthetic switch that activates when bound to its DNA target site. The system uses two CRISPR-Cas complexes to colocalize components of a de novo-designed protein switch (Co-LOCKR) to adjacent sites in the genome. Colocalization triggers a conformational change in the switch from an inactive closed state to an active open state with an exposed functional peptide. We prototype the system in yeast and demonstrate that DNA binding triggers activation of the switch, recruitment of a transcription factor, and expression of a downstream reporter gene. This DNA-triggered Co-LOCKR switch provides a platform to engineer sophisticated functions that should only be executed at a specific target site within the genome, with potential applications in a wide range of synthetic systems including epigenetic regulation, imaging, and genetic logic circuits.
journal_name
ACS Synth Bioljournal_title
ACS synthetic biologyauthors
Kirkpatrick RL,Lewis K,Langan RA,Lajoie MJ,Boyken SE,Eakman M,Baker D,Zalatan JGdoi
10.1021/acssynbio.0c00012subject
Has Abstractpub_date
2020-09-18 00:00:00pages
2316-2323issue
9issn
2161-5063journal_volume
9pub_type
杂志文章abstract：:We developed a hybrid synthetic circuit that co-opts the genetic regulation of the native bacterial quorum sensing autoinducer-2 and imposes an extra external controller for maintaining tightly controlled gene expression. This dual-input genetic controller was mathematically modeled and, by design, can be operated in ...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.0c00179
更新日期：2020-10-16 00:00:00
abstract：:Fucosyl-oligosaccharides (FOSs) play physiologically important roles as prebiotics, neuronal growth factors, and inhibitors of enteropathogens. However, challenges in designed synthesis and mass production of FOSs hamper their industrial applications. Here, we report flexible biosynthetic routes to produce various FOS...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.8b00436
更新日期：2019-02-15 00:00:00
abstract：:The Synthetic Biology Open Language (SBOL) is an emerging synthetic biology data exchange standard, designed primarily for unambiguous and efficient machine-to-machine communication. However, manual editing of SBOL is generally difficult for nontrivial designs. Here, we describe ShortBOL, a lightweight SBOL scripting ...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.9b00470
更新日期：2020-04-17 00:00:00
abstract：:A bacterial flavin transferase (ApbE) was recently employed for flavin mononucleotide (FMN) modification on the Na+-translocating NADH:quinone oxidoreductase C (NqrC) protein in the pathogenic Gram-negative bacterium Vibrio cholerae. We employed this unique post-translational modification in mammalian cells and found ...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.6b00284
更新日期：2017-04-21 00:00:00
abstract：:Multidrug resistance (MDR) is a globally relevant problem that requires novel approaches. Two-component systems are a promising, yet untapped target for novel antibacterials. They are prevalent in bacteria and absent in mammals, and their activity can be modulated upon perception of various stimuli. Screening pre-exis...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.6b00288
更新日期：2017-07-21 00:00:00
abstract：:Genome engineering technologies now enable precise manipulation of organism genotype, but can be limited in scalability by their design requirements. Here we describe Merlin ( http://merlincad.org ), an open-source web-based tool to assist biologists in designing experiments using multiplex automated genome engineerin...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.5b00219
更新日期：2016-06-17 00:00:00
abstract：:The extracellular expression of recombinant proteins using laboratory strains of Escherichia coli is now routinely achieved using naturally secreted substrates, such as YebF or the osmotically inducible protein Y (OsmY), as carrier molecules. However, secretion efficiency through these pathways needs to be improved fo...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.6b00366
更新日期：2017-05-19 00:00:00
abstract：:RNAs mediate many different processes that are central to cellular function. The ability to quantify or image RNAs in live cells is very useful in elucidating such functions of RNA. RNA aptamer-fluorogen systems have been increasingly used in labeling RNAs in live cells. Here, we use the malachite green aptamer (MGA),...
journal_title：ACS synthetic biology
pub_type： 信件
doi：10.1021/acssynbio.7b00237
更新日期：2018-03-16 00:00:00
abstract：:Cell-free protein synthesis (CFPS) is a widely used research tool in systems and synthetic biology. However, if CFPS is to become a mainstream technology for applications such as point of care manufacturing, we must understand the performance limits and costs of these systems. Toward this question, we used sequence sp...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.7b00465
更新日期：2018-08-17 00:00:00
abstract：:In many biotechnological applications, it is useful for gene expression to be regulated by multiple signals, as this allows the programming of complex behavior. Here we implement, in Escherichia coli, a system that compares the concentration of two signal molecules, and tunes GFP expression proportionally to their rel...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.7b00109
更新日期：2017-10-20 00:00:00
abstract：:Constructing efficient cellular factories often requires integration of heterologous pathways for synthesis of novel compounds and improved cellular productivity. Few genomic sites are routinely used, however, for efficient integration and expression of heterologous genes, especially in nonmodel hosts. Here, a data-gu...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.0c00299
更新日期：2020-09-18 00:00:00
abstract：:Lignans are plant secondary metabolites with a wide range of reported health-promoting bioactivities. Traditional routes toward these natural products involve, among others, the extraction from plant sources and chemical synthesis. However, the availability of the sources and the complex chemical structures of lignans...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.0c00354
更新日期：2020-11-20 00:00:00
abstract：:Metabolic engineering of Escherichia coli was performed to construct a 100% rationally engineered strain capable of overproducing L-isoleucine, an important branched-chain amino acid. The thrABC (encoding L-threonine biosynthetic enzymes), ilvA (encoding feedback-resistant threonine dehydratase), ilvIH (encoding feedb...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/sb300071a
更新日期：2012-11-16 00:00:00
abstract：:Light-switchable proteins enable unparalleled control of molecular biological processes in live organisms. Previously, we have engineered red/far-red and green/red photoreversible two-component signal transduction systems (TCSs) with transcriptional outputs in E. coli and used them to characterize and control syntheti...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/sb500273n
更新日期：2014-11-21 00:00:00
abstract：:The CRISPR-Cas9 nuclease has been repurposed as a tool for gene repression (CRISPRi). This catalytically dead Cas9 (dCas9) variant inhibits transcription by blocking either initiation or elongation by the RNA polymerase complex. Conditional control of dCas9-mediated repression has been achieved with inducible promoter...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.8b00347
更新日期：2019-02-15 00:00:00
abstract：:Strain engineering for synthetic biology and metabolic engineering applications often requires the expression of foreign proteins that can reduce cellular fitness. In order to quantify and visualize the evolutionary stability dynamics in engineered populations of Escherichia coli , we constructed randomized CMY (cyan-...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/sb400055h
更新日期：2013-09-20 00:00:00
abstract：:A major force shaping form and patterns in biology is based in the presence of amplification mechanisms able to generate ordered, large-scale spatial structures out of local interactions and random initial conditions. Turing patterns are one of the best known candidates for such ordering dynamics, and their existence ...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.5b00254
更新日期：2016-07-15 00:00:00
abstract：:Episodic gene expression, with periods of high expression separated by periods of no expression, is a pervasive biological phenomenon. This bursty pattern of expression draws from a finite reservoir of expression machinery in a highly time variant way, i.e., requiring no resources most of the time but drawing heavily ...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.6b00189
更新日期：2017-02-17 00:00:00
abstract：:Cell-free systems, as part of the synthetic biology field, have become a critical platform in 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 switc...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.0c00211
更新日期：2020-08-21 00:00:00
abstract：:We study translation repression in bacteria by engineering a regulatory circuit that functions as a binding assay for RNA binding proteins (RBP) in vivo. We do so by inducing expression of a fluorescent protein-RBP chimera, together with encoding its binding site at various positions within the ribosomal initiation re...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.8b00378
更新日期：2018-12-21 00:00:00
abstract：:Synthetic biology and metabolic engineering offer potentially green and attractive routes to the production of high value compounds. The provision of high-quality parts and pathways is crucial in enabling the biosynthesis of chemicals using synthetic biology. While a number of regulatory parts that provide control at ...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.0c00375
更新日期：2020-10-16 00:00:00
abstract：:Gene expression noise arises from stochastic variation in the synthesis and degradation of mRNA and protein molecules and creates differences in protein numbers across populations of genetically identical cells. Such variability can lead to imprecision and reduced performance of both native and synthetic networks. In ...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.8b00279
更新日期：2018-11-16 00:00:00
abstract：:Feedback allows biological systems to control gene expression precisely and reliably, even in the presence of uncertainty, by sensing and processing environmental changes. Taking inspiration from natural architectures, synthetic biologists have engineered feedback loops to tune the dynamics and improve the robustness ...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.8b00040
更新日期：2018-05-18 00:00:00
abstract：:An RNA-dependent RNA polymerase ribozyme that was highly optimized through in vitro evolution for the ability to copy a broad range of template sequences exhibits promiscuity toward other nucleic acids and nucleic acid analogues, including DNA, threose nucleic acid (TNA), and arabinose nucleic acid (ANA). By operating...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.9b00044
更新日期：2019-05-17 00:00:00
abstract：:A common approach to design genetic circuits is to compose gene expression cassettes together. While appealing, this modular approach is challenged by the fact that expression of each gene depends on the availability of transcriptional/translational resources, which is in turn determined by the presence of other genes...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.6b00361
更新日期：2017-07-21 00:00:00
abstract：:Synthetic biology is today harnessing the design of novel and greener biosynthesis routes for the production of added-value chemicals and natural products. The design of novel pathways often requires a detailed selection of enzyme sequences to import into the chassis at each of the reaction steps. To address such desi...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.5b00294
更新日期：2016-06-17 00:00:00
abstract：:A streamlined approach toward the rapid fabrication of streptavidin-biotin-based protein microarrays was investigated. First, using our engineered versatile plasmid (pBADcM-tBirA) and an optimal coexpression strategy for biotin ligase and biotin acceptor peptide (BAP) chimeric recombinant protein, an autogeneration sy...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.0c00343
更新日期：2020-09-18 00:00:00
abstract：:In this work, we develop methodologies for analyzing and cross comparing metabolic models. We investigate three important metabolic networks to discuss the complexity of biological organization of organisms, modeling, and system properties. In particular, we analyze these metabolic networks because of their biotechnol...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/sb300102k
更新日期：2013-05-17 00:00:00
abstract：:Crossbreeding is a classical yeast hybridization procedure, where the mating of haploid cells of opposite mating-type, MATa and MATα cells, produces a new heterozygous diploid. Here, we describe a method to generate haploid MATa and MATα cells using mating-type conversion caused by expression of the HO gene, which enc...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/sb400016j
更新日期：2013-12-20 00:00:00
abstract：:Fluorescent proteins are widely used as fusion tags to detect protein expression in vivo. To become fluorescent, these proteins must undergo chromophore maturation, a slow process with a half-time of 5 to >30 min that causes delays in real-time detection of protein expression. Here, we engineer a genetically encoded f...
journal_title：ACS synthetic biology
pub_type： 杂志文章
doi：10.1021/acssynbio.0c00407
更新日期：2020-11-20 00:00:00
