Riboswitches are highly conserved RNA elements that located in the 5’-UTR of m RNAs,which undergo real-time structure conformational change to achieve the regulation of downstream gene expression by sensing their cog...Riboswitches are highly conserved RNA elements that located in the 5’-UTR of m RNAs,which undergo real-time structure conformational change to achieve the regulation of downstream gene expression by sensing their cognate ligands.S-adenosylmethionine(SAM)is a ubiquitous methyl donor for transmethylation reactions in all living organisms.SAM riboswitch is one of the most abundant riboswitches that bind to SAM with high affinity and selectivity,serving as regulatory modules in multiple metabolic pathways.To date,seven SAM-specific riboswitch classes that belong to four families,one SAM/SAH riboswitch and one SAH riboswitch have been identified.Each SAM riboswitch family has a well-organized tertiary core scaffold to support their unique ligand-specific binding pocket.In this review,we summarize the current research progress on the distribution,structure,ligand recognition and gene regulation mechanism of these SAM-related riboswitch families,and further discuss their evolutionary prospects and potential applications.展开更多
Riboswitches are functional RNA elements that regulate gene expression by directly detecting metabolites.Twenty years have passed since it was first discovered,researches on riboswitches are becoming increasingly stan...Riboswitches are functional RNA elements that regulate gene expression by directly detecting metabolites.Twenty years have passed since it was first discovered,researches on riboswitches are becoming increasingly standardized and refined,which could significantly promote people’s cognition of RNA function as well.Here,we focus on some representative orphan riboswitches,enumerate the structural and functional transformation and artificial design of riboswitches including the coupling with ribozymes,hoping to attain a comprehensive understanding of riboswitch research.展开更多
With the support by the National Natural Science Foundation of China,the research team led by Dr.Liu Yuchen(刘宇辰)at the State Engineering Laboratory of Medical Key Technologies Application of Synthetic Biology,Shenz...With the support by the National Natural Science Foundation of China,the research team led by Dr.Liu Yuchen(刘宇辰)at the State Engineering Laboratory of Medical Key Technologies Application of Synthetic Biology,Shenzhen Second People’s Hospital,the First Affiliated Hospital of Shenzhen University,developed a type of CRISPR-based riboswitch,which was published in Nature Methods(2016,doi:10.展开更多
Most riboswitches are characterized by two components, an aptamer domain that folds into a unique ligand binding pocket to interact with the ligand, and an expression platform that converts folding changes in the apta...Most riboswitches are characterized by two components, an aptamer domain that folds into a unique ligand binding pocket to interact with the ligand, and an expression platform that converts folding changes in the aptamer into changes in gene expression. Using the recently developed systematic helix-based computational method, we theoretically studied the refolding and co-transcriptional folding behaviors of the purine riboswitch aptamers from Bacillus subtilis xpt-pbu X guanine riboswitch and Vibrio vulnificus add adenine riboswitch. Despite several intermediate structures persisting a short time during the transcription, helices P2, P3 and P1 fold in turn for both aptamers. Although some misfolded structures are observed during the refolding process, the RNAs can fold into the ligand binding pocket structure containing helices P2, P3 and P1 within a few seconds, suggesting the aptamer domains are highly evolved. The purine riboswitch aptamers can quickly fold into the ligand binding pocket structure even at a high transcription speed, possibly because formation of this structure is the necessary prerequisite for the riboswitch to bind its ligand and then regulate relevant gene expression.展开更多
RNAs carry out diverse biological functions, partly because different conformations of the same RNA sequence can play different roles in cellular activities. To fully understand the biological functions of RNAs requir...RNAs carry out diverse biological functions, partly because different conformations of the same RNA sequence can play different roles in cellular activities. To fully understand the biological functions of RNAs requires a conceptual framework to investigate the folding kinetics of RNA molecules, instead of native structures alone. Over the past several decades, many experimental and theoretical methods have been developed to address RNA folding. The helix-based RNA folding theory is the one which uses helices as building blocks, to calculate folding kinetics of secondary structures with pseudoknots of long RNA in two different folding scenarios. Here, we will briefly review the helix-based RNA folding theory and its application in exploring regulation mechanisms of several riboswitches and self-cleavage activities of the hepatitis delta virus (HDV) ribozyme.展开更多
T box sequences have been identified upstream of a large number of uncharacterized genes such as transporters in bacterial genomes. Expression of each T box family gene is induced by limitation for a specific amino ac...T box sequences have been identified upstream of a large number of uncharacterized genes such as transporters in bacterial genomes. Expression of each T box family gene is induced by limitation for a specific amino acid. T box family genes contain an untranslated leader region containing a factor-independent transcriptional terminator upstream of the structural genes. The anticodon of uncharged tRNA base-pairs with the leader mRNA at a codon referred to as the specifier sequence, inducing formation of an alternative antiterminator structure, allowing expression of the structural genes. There are several additional conserved primary sequence and secondary structural elements. Analysis of these elements can be used to predict the identity of the specifier codon and the amino acid signal.?Bacillus subtilis hypothetical amino acid permease, yvbW, was analyzed as an example of this type of transcriptional regulatory prediction suggesting expression in response to leucine limitation. Expression was induced up to 130-fold in response to leucine limitation, utilizing a yvbW-lacZ transcriptional fusion. These data suggest that hypothetical amino acid permease YvbW may participate in leucine metabolism. A yvbW knockout strain was generated, although the substrate specificity for the putative amino acid permease was not identified.展开更多
Riboswitches are conserved RNA elements that specifically recognize the cognate metabolites and regulate downstream gene expression involved in the metabolic pathways.To date,two classes of xanthine-responsive riboswi...Riboswitches are conserved RNA elements that specifically recognize the cognate metabolites and regulate downstream gene expression involved in the metabolic pathways.To date,two classes of xanthine-responsive riboswitches involved in xanthine homeostasis have been identified.The recently reported xanthine-II riboswitch originates from guanine riboswitch family,featuring a single U-to-G mutation and several nucleotide insertions.Here,we report the complex structure of xanthine-II riboswitch bound to xanthine.The tertiary structure of xanthine-II riboswitch adopts a three-way junction scaffold similar to that of guanine riboswitch.However,the distinctive mutation and insertions in xanthine-II riboswitch facilitate the formation of a highly specific binding pocket for xanthine,distinguishing it from guanine riboswitches.Xanthine is bound in the junction region,forming a base triple with C64 and the mutant nucleotide G37,and is sandwiched by one base pair U8-A38 and one base triple A7-C36-U65.Structural alignment and ligand recognition specificity of the xanthine-II riboswitch are further verified by ligand-binding assays of structure-based mutation using isothermal titration calorimetry.Furthermore,leveraging the ligand specificity of the xanthine-II riboswitch,we develop a highly specific and sensitive biosensor for xanthine detection by fusing xanthine-II riboswitch with Pepper fluorogenic aptamer,highlighting the potential applications of xanthine-II riboswitch in diagnosing diseases related to xanthine metabolism disorders.展开更多
Bacteria exhibit a rich repertoire of RNA molecules that intricately regulate gene expression at multiple hierarchical levels,including small RNAs(sRNAs),riboswitches,and antisense RNAs.Notably,the majority of these r...Bacteria exhibit a rich repertoire of RNA molecules that intricately regulate gene expression at multiple hierarchical levels,including small RNAs(sRNAs),riboswitches,and antisense RNAs.Notably,the majority of these regulatory RNAs lack or have limited protein-coding capacity but play pivotal roles in orchestrating gene expression by modulating transcription,post-transcription or translation processes.Leveraging and redesigning these regulatory RNA elements have emerged as pivotal strategies in the domains of metabolic engineering and synthetic biology.While previous investigations predominantly focused on delineating the roles of regulatory RNA in Gram-negative bacterial models such as Escherichia coli and Salmonella enterica,this review aims to summarize the mechanisms and functionalities of endogenous regulatory RNAs inherent to typical Gram-positive bacteria,notably Bacillus subtilis.Furthermore,we explore the engineering and practical applications of these regulatory RNA elements in the arena of synthetic biology,employing B.subtilis as a foundational chassis.展开更多
Genetically encoded circuits have been successfully utilized to assess and characterize target variants with desirable traits from large mutant libraries.Adenosylcobalamin is an essential coenzyme that is required in ...Genetically encoded circuits have been successfully utilized to assess and characterize target variants with desirable traits from large mutant libraries.Adenosylcobalamin is an essential coenzyme that is required in many intracellular physiological reactions and is widely used in the pharmaceutical and food industries.High-throughput screening techniques capable of detecting adenosylcobalamin productivity and selecting superior adenosylcobalamin biosynthesis strains are critical for the creation of an effective microbial cell factory for the production of adenosylcobalamin at an industrial level.In this study,we developed an RNA-protein hybrid biosensor whose input part was an endogenous RNA riboswitch to specifically respond to adenosylcobalamin,the inverter part was an orthogonal transcriptional repressor to obtain signal inversion,and the output part was a fluorescent protein to be easily detected.The hybrid biosensor could specifically and positively correlate adenosylcobalamin concentrations to green fluorescent protein expression levels in vivo.This study also improved the operating concentration and dynamic range of the hybrid biosensor by systematic optimization.An individual cell harboring the hybrid biosensor presented over 20-fold higher fluorescence intensity than the negative control.Then,using such a biosensor combined with fluorescence-activated cell sorting,we established a high-throughput screening platform for screening adenosylcobalamin overproducers.This study demonstrates that this platform has significant potential to quickly isolate high-productive strains to meet industrial demand and that the framework is acceptable for various metabolites.展开更多
Since nucleic acids(DNA and RNA) play very important roles in cells,they are molecular targets of many clinically used drugs,such as anticancer drugs and antibiotics.Because of clinical demands for treating various de...Since nucleic acids(DNA and RNA) play very important roles in cells,they are molecular targets of many clinically used drugs,such as anticancer drugs and antibiotics.Because of clinical demands for treating various deadly cancers and drug-resistant strains of pathogens,there are urgent needs to develop novel therapeutic agents.Targeting nucleic acids hasn’t been the mainstream of drug discovery in the past,and the lack of 3D structural information for designing and developing drug specificity is one of the main reasons.Fortunately,many important structures of nucleic acids and their protein complexes have been determined over the past decade,which provide novel platforms for future drug design and discovery.In this review,we describe some useful nucleic acid structures,particularly their interactions with the ligands and therapeutic candidates or even drugs.We summarize important information for designing novel potent drugs and for targeting nucleic acids and protein-nucleic acid complexes to treat cancers and overcome the drug-resistant problems.展开更多
Overproduction of small-molecule chemicals using engineered microbial cells has greatly reduced the production cost and promoted environmental protection. Notably, the rapid and sensitive evaluation of the in vivo con...Overproduction of small-molecule chemicals using engineered microbial cells has greatly reduced the production cost and promoted environmental protection. Notably, the rapid and sensitive evaluation of the in vivo concentrations of the desired products greatly facilitates the optimization process of cell factories. For this purpose, many genetic components have been adapted into in vivo biosensors of small molecules, which couple the intracel- lular concentrations of small molecules to easily detectable readouts such as fluorescence, absorbance, and cell growth. Such biosensors allow a high-throughput screen- ing of the small-molecule products, and can be roughly classified as protein-based and RNA-based biosensors. This review summarizes the recent developments in the design and applications of biosensors for small-molecule products.展开更多
A parallel screening of 27 different flavonoids and chalcones was conducted using 6 artificial naringenin-activated riboswitches(M1,M2,M3,O,L and H).A quantitative structure-property relationship approach was applied ...A parallel screening of 27 different flavonoids and chalcones was conducted using 6 artificial naringenin-activated riboswitches(M1,M2,M3,O,L and H).A quantitative structure-property relationship approach was applied to understand the physicochemical properties of the flavonoid structures resulting in specificity differences relied on the fluorescence intensity of a green fluorescent protein reporter.Robust models of riboswitches M1,M2 and O that had good predictive power were constructed with descriptors selected for their high correlation.Increased electronegativity and hydrophilicity of the flavonoids structures were identified as two properties that increased binding affinity to RNA riboswitches.Hydroxyl groups at the C-3′and C-4’positions of the flavonoid molecule were strictly required for ligand-activation with riboswitches M1 and M2.Riboswitches O and L preferred multi-hydroxylated flavones as ligands.Substitutions on the A ring of the flavonoid molecule were not important in the molecular recognition process.O-glycosylated derivatives were not recognized by any of the riboswitches,presumably due to steric hindrances.Despite the challenges of detecting RNA conformational change after ligand binding,the resulting models elucidate important physicochemical features in the ligands for conformational structural studies of artificial aptamer complexes and for design of ligands having higher binding specificity.展开更多
Transcription attenuation in response to the availability of a specific amino acid is believed to be controlled by alternative configurations of RNA secondary structures that lead to the arrest of translation or the r...Transcription attenuation in response to the availability of a specific amino acid is believed to be controlled by alternative configurations of RNA secondary structures that lead to the arrest of translation or the release of the arrested ribosome from the leader mRNA molecule.In this study,we first report a possible example of the DnaA‐dependent riboswitch for transcription attenuation in Escherichia coli.We show that(i)DnaA regulates the transcription of the structural genes but not that of the leader hisL gene;(ii)DnaA might bind to rDnaA boxes present in the HisL‐SL RNA,and subsequently attenuate the transcription of the operon;(iii)the HisL‐SL RNA and rDnaA boxes are phylogenetically conserved and evolutionarily important;and(iv)the translating ribosome is required for deattenuation of the his operon,whereas tRNA^(His) strengthens attenuation.This mechanism seems to be phylogenetically conserved in Gram‐negative bacteria and evolutionarily important.展开更多
基金supported by the National Natural Science Foundation of China(32022039,31870810,91940302,91640104)the National Key Research and Development Project of China(2021YFC2300300)+2 种基金the China Postdoctoral Science Foundation(2022M713637)the Outstanding Youth Fund of Zhejiang Province(LR19C050003)the Fundamental Research Funds for the Central Universities(2017QN81010)。
文摘Riboswitches are highly conserved RNA elements that located in the 5’-UTR of m RNAs,which undergo real-time structure conformational change to achieve the regulation of downstream gene expression by sensing their cognate ligands.S-adenosylmethionine(SAM)is a ubiquitous methyl donor for transmethylation reactions in all living organisms.SAM riboswitch is one of the most abundant riboswitches that bind to SAM with high affinity and selectivity,serving as regulatory modules in multiple metabolic pathways.To date,seven SAM-specific riboswitch classes that belong to four families,one SAM/SAH riboswitch and one SAH riboswitch have been identified.Each SAM riboswitch family has a well-organized tertiary core scaffold to support their unique ligand-specific binding pocket.In this review,we summarize the current research progress on the distribution,structure,ligand recognition and gene regulation mechanism of these SAM-related riboswitch families,and further discuss their evolutionary prospects and potential applications.
基金the National Key Research and Development Program of China(2021YFC2100700)the National Natural Science Foundation of China(Grant NSFC-22278313).
文摘Riboswitches are functional RNA elements that regulate gene expression by directly detecting metabolites.Twenty years have passed since it was first discovered,researches on riboswitches are becoming increasingly standardized and refined,which could significantly promote people’s cognition of RNA function as well.Here,we focus on some representative orphan riboswitches,enumerate the structural and functional transformation and artificial design of riboswitches including the coupling with ribozymes,hoping to attain a comprehensive understanding of riboswitch research.
文摘With the support by the National Natural Science Foundation of China,the research team led by Dr.Liu Yuchen(刘宇辰)at the State Engineering Laboratory of Medical Key Technologies Application of Synthetic Biology,Shenzhen Second People’s Hospital,the First Affiliated Hospital of Shenzhen University,developed a type of CRISPR-based riboswitch,which was published in Nature Methods(2016,doi:10.
基金Supported by the National Natural Science Foundation of China(31600592)Hubei Key Laboratory of Economic Forest Germplasm Improvement and Resources Comprehensive Utilization,Hubei Collaborative Innovation Center for the Characteristic Resources Exploitation of Dabie Mountains(2017BX08)
文摘Most riboswitches are characterized by two components, an aptamer domain that folds into a unique ligand binding pocket to interact with the ligand, and an expression platform that converts folding changes in the aptamer into changes in gene expression. Using the recently developed systematic helix-based computational method, we theoretically studied the refolding and co-transcriptional folding behaviors of the purine riboswitch aptamers from Bacillus subtilis xpt-pbu X guanine riboswitch and Vibrio vulnificus add adenine riboswitch. Despite several intermediate structures persisting a short time during the transcription, helices P2, P3 and P1 fold in turn for both aptamers. Although some misfolded structures are observed during the refolding process, the RNAs can fold into the ligand binding pocket structure containing helices P2, P3 and P1 within a few seconds, suggesting the aptamer domains are highly evolved. The purine riboswitch aptamers can quickly fold into the ligand binding pocket structure even at a high transcription speed, possibly because formation of this structure is the necessary prerequisite for the riboswitch to bind its ligand and then regulate relevant gene expression.
基金Project supported by the Science Fund from the Key Laboratory of Hubei Province, China (Grant No. 201932003)the National Natural Science Foundation of China (Grant Nos. 1157324 and 31600592).
文摘RNAs carry out diverse biological functions, partly because different conformations of the same RNA sequence can play different roles in cellular activities. To fully understand the biological functions of RNAs requires a conceptual framework to investigate the folding kinetics of RNA molecules, instead of native structures alone. Over the past several decades, many experimental and theoretical methods have been developed to address RNA folding. The helix-based RNA folding theory is the one which uses helices as building blocks, to calculate folding kinetics of secondary structures with pseudoknots of long RNA in two different folding scenarios. Here, we will briefly review the helix-based RNA folding theory and its application in exploring regulation mechanisms of several riboswitches and self-cleavage activities of the hepatitis delta virus (HDV) ribozyme.
文摘T box sequences have been identified upstream of a large number of uncharacterized genes such as transporters in bacterial genomes. Expression of each T box family gene is induced by limitation for a specific amino acid. T box family genes contain an untranslated leader region containing a factor-independent transcriptional terminator upstream of the structural genes. The anticodon of uncharged tRNA base-pairs with the leader mRNA at a codon referred to as the specifier sequence, inducing formation of an alternative antiterminator structure, allowing expression of the structural genes. There are several additional conserved primary sequence and secondary structural elements. Analysis of these elements can be used to predict the identity of the specifier codon and the amino acid signal.?Bacillus subtilis hypothetical amino acid permease, yvbW, was analyzed as an example of this type of transcriptional regulatory prediction suggesting expression in response to leucine limitation. Expression was induced up to 130-fold in response to leucine limitation, utilizing a yvbW-lacZ transcriptional fusion. These data suggest that hypothetical amino acid permease YvbW may participate in leucine metabolism. A yvbW knockout strain was generated, although the substrate specificity for the putative amino acid permease was not identified.
基金supported by the National Key Research and Development Project of China (2021YFC2300300,2023YFC2604300)the National Natural Science Foundation of China (32325029,91940302,91640104)。
文摘Riboswitches are conserved RNA elements that specifically recognize the cognate metabolites and regulate downstream gene expression involved in the metabolic pathways.To date,two classes of xanthine-responsive riboswitches involved in xanthine homeostasis have been identified.The recently reported xanthine-II riboswitch originates from guanine riboswitch family,featuring a single U-to-G mutation and several nucleotide insertions.Here,we report the complex structure of xanthine-II riboswitch bound to xanthine.The tertiary structure of xanthine-II riboswitch adopts a three-way junction scaffold similar to that of guanine riboswitch.However,the distinctive mutation and insertions in xanthine-II riboswitch facilitate the formation of a highly specific binding pocket for xanthine,distinguishing it from guanine riboswitches.Xanthine is bound in the junction region,forming a base triple with C64 and the mutant nucleotide G37,and is sandwiched by one base pair U8-A38 and one base triple A7-C36-U65.Structural alignment and ligand recognition specificity of the xanthine-II riboswitch are further verified by ligand-binding assays of structure-based mutation using isothermal titration calorimetry.Furthermore,leveraging the ligand specificity of the xanthine-II riboswitch,we develop a highly specific and sensitive biosensor for xanthine detection by fusing xanthine-II riboswitch with Pepper fluorogenic aptamer,highlighting the potential applications of xanthine-II riboswitch in diagnosing diseases related to xanthine metabolism disorders.
基金This work was supported by the National Natural Science Foundation of China(31970085 and 32000058)the National Key Research and Development Program of China(2021YFC2100800).
文摘Bacteria exhibit a rich repertoire of RNA molecules that intricately regulate gene expression at multiple hierarchical levels,including small RNAs(sRNAs),riboswitches,and antisense RNAs.Notably,the majority of these regulatory RNAs lack or have limited protein-coding capacity but play pivotal roles in orchestrating gene expression by modulating transcription,post-transcription or translation processes.Leveraging and redesigning these regulatory RNA elements have emerged as pivotal strategies in the domains of metabolic engineering and synthetic biology.While previous investigations predominantly focused on delineating the roles of regulatory RNA in Gram-negative bacterial models such as Escherichia coli and Salmonella enterica,this review aims to summarize the mechanisms and functionalities of endogenous regulatory RNAs inherent to typical Gram-positive bacteria,notably Bacillus subtilis.Furthermore,we explore the engineering and practical applications of these regulatory RNA elements in the arena of synthetic biology,employing B.subtilis as a foundational chassis.
基金supported by the National Key R&D Program of China[Grant number 2021YFC2100700]the National Natural Science Foundation of China[Grant numbers 22178372,32300069]+3 种基金TIB-VIB Joint Center of Synthetic Biology[Grant number TSBICIP-IJCP-002]the National Science Fund for Distinguished Young Scholars[Grant number 22325807]the Tianjin Synthetic Biotechnology Innovation Capacity Improvement Project[Grant numbers TSBICIP-KJGG-011,TSBICIP-CXRC-055]the Yellow River Delta Industry Leading Talents[Grant number DYRC20190212].
文摘Genetically encoded circuits have been successfully utilized to assess and characterize target variants with desirable traits from large mutant libraries.Adenosylcobalamin is an essential coenzyme that is required in many intracellular physiological reactions and is widely used in the pharmaceutical and food industries.High-throughput screening techniques capable of detecting adenosylcobalamin productivity and selecting superior adenosylcobalamin biosynthesis strains are critical for the creation of an effective microbial cell factory for the production of adenosylcobalamin at an industrial level.In this study,we developed an RNA-protein hybrid biosensor whose input part was an endogenous RNA riboswitch to specifically respond to adenosylcobalamin,the inverter part was an orthogonal transcriptional repressor to obtain signal inversion,and the output part was a fluorescent protein to be easily detected.The hybrid biosensor could specifically and positively correlate adenosylcobalamin concentrations to green fluorescent protein expression levels in vivo.This study also improved the operating concentration and dynamic range of the hybrid biosensor by systematic optimization.An individual cell harboring the hybrid biosensor presented over 20-fold higher fluorescence intensity than the negative control.Then,using such a biosensor combined with fluorescence-activated cell sorting,we established a high-throughput screening platform for screening adenosylcobalamin overproducers.This study demonstrates that this platform has significant potential to quickly isolate high-productive strains to meet industrial demand and that the framework is acceptable for various metabolites.
基金financially supported by the Georgia Cancer Coalition(GCC) Distinguished Cancer Clinicians and Scientists and by the US National Science Foundation(NSF MCB-0824837)
文摘Since nucleic acids(DNA and RNA) play very important roles in cells,they are molecular targets of many clinically used drugs,such as anticancer drugs and antibiotics.Because of clinical demands for treating various deadly cancers and drug-resistant strains of pathogens,there are urgent needs to develop novel therapeutic agents.Targeting nucleic acids hasn’t been the mainstream of drug discovery in the past,and the lack of 3D structural information for designing and developing drug specificity is one of the main reasons.Fortunately,many important structures of nucleic acids and their protein complexes have been determined over the past decade,which provide novel platforms for future drug design and discovery.In this review,we describe some useful nucleic acid structures,particularly their interactions with the ligands and therapeutic candidates or even drugs.We summarize important information for designing novel potent drugs and for targeting nucleic acids and protein-nucleic acid complexes to treat cancers and overcome the drug-resistant problems.
文摘Overproduction of small-molecule chemicals using engineered microbial cells has greatly reduced the production cost and promoted environmental protection. Notably, the rapid and sensitive evaluation of the in vivo concentrations of the desired products greatly facilitates the optimization process of cell factories. For this purpose, many genetic components have been adapted into in vivo biosensors of small molecules, which couple the intracel- lular concentrations of small molecules to easily detectable readouts such as fluorescence, absorbance, and cell growth. Such biosensors allow a high-throughput screen- ing of the small-molecule products, and can be roughly classified as protein-based and RNA-based biosensors. This review summarizes the recent developments in the design and applications of biosensors for small-molecule products.
基金This work was supported by the Global Research Laboratory Program(NRF 2016K1A1A2912829)through the National Research Foundation of Korea(NRF)This work was also supported by the National Natural Science Foundation of China(grant number 21636001)+3 种基金Research Program of Beijing Municipal Education Commission(grant number KM201911417012)This work was also financially supported by the Office of the Ministry of Higher Education,Science,Research and Innovationthe Thailand Science Research and Innovation through the Kasetsart University Reinventing University Program 2021 and Department of Zoology,Faculty of Science,Kasetsart UniversityP.P.would like to acknowledge the supports provided by the International Affairs under the scholarship program of ASEAN+6 and Interdisciplinary Graduate Program in Bioscience in Kasetsart University.
文摘A parallel screening of 27 different flavonoids and chalcones was conducted using 6 artificial naringenin-activated riboswitches(M1,M2,M3,O,L and H).A quantitative structure-property relationship approach was applied to understand the physicochemical properties of the flavonoid structures resulting in specificity differences relied on the fluorescence intensity of a green fluorescent protein reporter.Robust models of riboswitches M1,M2 and O that had good predictive power were constructed with descriptors selected for their high correlation.Increased electronegativity and hydrophilicity of the flavonoids structures were identified as two properties that increased binding affinity to RNA riboswitches.Hydroxyl groups at the C-3′and C-4’positions of the flavonoid molecule were strictly required for ligand-activation with riboswitches M1 and M2.Riboswitches O and L preferred multi-hydroxylated flavones as ligands.Substitutions on the A ring of the flavonoid molecule were not important in the molecular recognition process.O-glycosylated derivatives were not recognized by any of the riboswitches,presumably due to steric hindrances.Despite the challenges of detecting RNA conformational change after ligand binding,the resulting models elucidate important physicochemical features in the ligands for conformational structural studies of artificial aptamer complexes and for design of ligands having higher binding specificity.
基金supported by grants from the National Natural Science Foundation of China(Grant no.32260233 to Morigen)the Science and Technology Foundation of Inner Mongolia(Inner Mongolia Key Laboratory for Molecular Regulation of the Cell,Grant no.2021PT0002).
文摘Transcription attenuation in response to the availability of a specific amino acid is believed to be controlled by alternative configurations of RNA secondary structures that lead to the arrest of translation or the release of the arrested ribosome from the leader mRNA molecule.In this study,we first report a possible example of the DnaA‐dependent riboswitch for transcription attenuation in Escherichia coli.We show that(i)DnaA regulates the transcription of the structural genes but not that of the leader hisL gene;(ii)DnaA might bind to rDnaA boxes present in the HisL‐SL RNA,and subsequently attenuate the transcription of the operon;(iii)the HisL‐SL RNA and rDnaA boxes are phylogenetically conserved and evolutionarily important;and(iv)the translating ribosome is required for deattenuation of the his operon,whereas tRNA^(His) strengthens attenuation.This mechanism seems to be phylogenetically conserved in Gram‐negative bacteria and evolutionarily important.
