Artificial synthesis and site-specific modification of peptides and proteins has evolved into an indispensable tool for protein engineers and chemical biologists. Chemical and enzymatic approaches to peptide ligation ...Artificial synthesis and site-specific modification of peptides and proteins has evolved into an indispensable tool for protein engineers and chemical biologists. Chemical and enzymatic approaches to peptide ligation are important alternatives of recombinant DNA technology for protein synthesis and modification. Although as old as that of chemical procedures, enzyme-mediated peptide ligation is far less developed than that of chemical counterpart due to the difficult availability of peptide ligase.Fortunately, this situation has been changed slowly with the fast development of biological techniques. In the past decades, several natural peptide ligases have been discovered. Protein engineering to improve the ligation efficiencies of the natural peptide ligase and to reverse the functionality of protease provide more powerful peptide ligases. In this review, the advances of enzyme-mediated peptide ligation and their application in protein synthesis and modification will be discussed.展开更多
Restriction enzyme-mediated integration(REMI) of DNA has been recently received attention as a new technique for the generation of mutants by transformation in fungi. Trichoderma atroviride strain T23 was transformed ...Restriction enzyme-mediated integration(REMI) of DNA has been recently received attention as a new technique for the generation of mutants by transformation in fungi. Trichoderma atroviride strain T23 was transformed with linearized plasmid pV2, conferring resistance to hygromycin B, in the presence of restriction enzyme used to linearize the plasmid. A total of 172 regeneration transformants were detected by successive inoculation for seven times subcultivation on fresh PDA plate containing hygromycin B. The plasmid was integrated stably into the chromosome DNA, which was confirmed by PCR and southern analysis. The difference between 172 transformants and the parent strain was confirmed in colonial color, sporulation and growth rate. The results showed that the significant difference appeared in above mentioned characters between transformants and parent strain is sporulation capability. Transformants TC6, TD5, TE7, TF1 and TK1 produced higher amounts of conidia than the parent strain T23. In addition, transformants TK1and TC6 showed stronger inhibition to the growth rate of the cucumber wilt pathogen (Fusarium oxyporum) in vitro.展开更多
Recent years have seen an ever increasing number of enzyme mediated protein/peptide modification reactions, which contribute significantly to the elucidation of related biological functions. The many available enzymes...Recent years have seen an ever increasing number of enzyme mediated protein/peptide modification reactions, which contribute significantly to the elucidation of related biological functions. The many available enzymes have, however, caused difficulties for practitioners in choosing the most appropriate enzyme for a certain purpose. This review surveyed the widely used enzymes(i.e., sortases, butelase 1,subtiligase, formylglycine generating enzyme and farnesyltransferase) in the manipulation of proteins/peptides, and the application fields of these enzymes as well as the advantages and limitations of each enzyme are summarized.展开更多
Site-specific protein labeling plays important roles in drug discovery and illuminating biological processes at the molecular level.However,it is challenging to label proteins with high specificity while not affecting...Site-specific protein labeling plays important roles in drug discovery and illuminating biological processes at the molecular level.However,it is challenging to label proteins with high specificity while not affecting their structures and biochemical activities.Over the last few years,a variety of promising strategies have been devised that address these challenges including those that involve introduction of small-size peptide tags or unnatural amino acids(UAAs),chemical labeling of specific protein residues,and affinity-driven labeling.This review summarizes recent developments made in the area of site-specific protein labeling utilizing genetically encoding-and chemical-based methods,and discusses future issues that need to be addressed by researchers in this field.展开更多
Uronic acids are prevalent components of crucial glycoconjugates,pivotal in various biological processes.In nature,NDP-uronic acids,the nucleosides-activated uronic acids,serve as glycosylation donors catalyzed by uro...Uronic acids are prevalent components of crucial glycoconjugates,pivotal in various biological processes.In nature,NDP-uronic acids,the nucleosides-activated uronic acids,serve as glycosylation donors catalyzed by uronosyltransferases(UATs)to construct glycans containing uronic acids.Despite their biological importance,the synthesis of naturally occurring NDP-uronic acids on a large scale remains challenging.Here,we developed an oxidation reaction insertion strategy for the efficient synthesis of NDP-uronic acids,and 11 NDP-uronic acids were successfully prepared in good yield and on a large scale.The prepared NDP-uronic acids can be used to explore new uronosyltransferases and synthesize uronic acids containing carbohydrates for fundamental research.展开更多
Molecularly imprinted polymers(MIPs)exhibit high selectivity resulting from imprinted cavities and superior performance from functional materials,which have attracted much attention in many fields.However,the combinat...Molecularly imprinted polymers(MIPs)exhibit high selectivity resulting from imprinted cavities and superior performance from functional materials,which have attracted much attention in many fields.However,the combination of MIPs film and functional materials is a great challenge.In this study,hemin/graphene hybrid nanosheets(H-GNs)were used to initiate the imprinted polymerization by catalyzing the generation of free radicals.Thus,MIPs using sulfamethoxazole as the template was directly prepared on the surface of H-GNs without any film modification.Most importantly,the template could be absorbed on the H-GNs to enhance the number of imprinted sites per unit surface area,which could improve the selectivity of MIPs film.Thus,the composites could exhibit high adsorption capacity(29.4 mg/g),imprinting factor(4.2)and excellent conductivity,which were modified on the surface of electrode for rapid,selective and sensitive detection of sulfamethoxazole in food and serum samples.The linear range was changed from 5μg/kg to 1 mg/g and the limit of detection was 1.2μg/kg.This sensor was free from interference caused by analogues of sulfamethoxazole,which provides a novel insight for the preparation of MIPs-based sensor and its application in food safety monitoring and human exposure study.展开更多
基金The National Natural Science Foundation of China (Nos. 21462023, 21778025)the Education Department of Jiangxi Province (No.150297)
文摘Artificial synthesis and site-specific modification of peptides and proteins has evolved into an indispensable tool for protein engineers and chemical biologists. Chemical and enzymatic approaches to peptide ligation are important alternatives of recombinant DNA technology for protein synthesis and modification. Although as old as that of chemical procedures, enzyme-mediated peptide ligation is far less developed than that of chemical counterpart due to the difficult availability of peptide ligase.Fortunately, this situation has been changed slowly with the fast development of biological techniques. In the past decades, several natural peptide ligases have been discovered. Protein engineering to improve the ligation efficiencies of the natural peptide ligase and to reverse the functionality of protease provide more powerful peptide ligases. In this review, the advances of enzyme-mediated peptide ligation and their application in protein synthesis and modification will be discussed.
基金Item supported by the Tenth Five-yearProgram of science and technology for creation of biocontrol a-gent against corn disease(2004BA509B0405)
文摘Restriction enzyme-mediated integration(REMI) of DNA has been recently received attention as a new technique for the generation of mutants by transformation in fungi. Trichoderma atroviride strain T23 was transformed with linearized plasmid pV2, conferring resistance to hygromycin B, in the presence of restriction enzyme used to linearize the plasmid. A total of 172 regeneration transformants were detected by successive inoculation for seven times subcultivation on fresh PDA plate containing hygromycin B. The plasmid was integrated stably into the chromosome DNA, which was confirmed by PCR and southern analysis. The difference between 172 transformants and the parent strain was confirmed in colonial color, sporulation and growth rate. The results showed that the significant difference appeared in above mentioned characters between transformants and parent strain is sporulation capability. Transformants TC6, TD5, TE7, TF1 and TK1 produced higher amounts of conidia than the parent strain T23. In addition, transformants TK1and TC6 showed stronger inhibition to the growth rate of the cucumber wilt pathogen (Fusarium oxyporum) in vitro.
基金The financial support from the National Recruitment Program of Global Youth Experts(1000 Talents Plan)the National Natural Science Foundation of China (No. 81703406)
文摘Recent years have seen an ever increasing number of enzyme mediated protein/peptide modification reactions, which contribute significantly to the elucidation of related biological functions. The many available enzymes have, however, caused difficulties for practitioners in choosing the most appropriate enzyme for a certain purpose. This review surveyed the widely used enzymes(i.e., sortases, butelase 1,subtiligase, formylglycine generating enzyme and farnesyltransferase) in the manipulation of proteins/peptides, and the application fields of these enzymes as well as the advantages and limitations of each enzyme are summarized.
基金supported by the National Key R&D Program of China(No.2021YFC2103600)the National Natural Science Foundation of China(Nos.22278224,22478191)+1 种基金the Project of Priority Academic Program Development of Jiangsu Higher Education Institutions(PAPD)the State Key Laboratory of Materials Oriented Chemical Engineering(No.KL21-08)。
文摘Site-specific protein labeling plays important roles in drug discovery and illuminating biological processes at the molecular level.However,it is challenging to label proteins with high specificity while not affecting their structures and biochemical activities.Over the last few years,a variety of promising strategies have been devised that address these challenges including those that involve introduction of small-size peptide tags or unnatural amino acids(UAAs),chemical labeling of specific protein residues,and affinity-driven labeling.This review summarizes recent developments made in the area of site-specific protein labeling utilizing genetically encoding-and chemical-based methods,and discusses future issues that need to be addressed by researchers in this field.
基金financially supported by National Natural Science Foundation of China(No.22207113 to J.Zhang)Guangdong Basic and Applied Basic Research Foundation(No.2021A1515110588to J.Zhang)Natural Science Foundation of Shanghai Municipality(No.22ZR1474000 to L.Wen)。
文摘Uronic acids are prevalent components of crucial glycoconjugates,pivotal in various biological processes.In nature,NDP-uronic acids,the nucleosides-activated uronic acids,serve as glycosylation donors catalyzed by uronosyltransferases(UATs)to construct glycans containing uronic acids.Despite their biological importance,the synthesis of naturally occurring NDP-uronic acids on a large scale remains challenging.Here,we developed an oxidation reaction insertion strategy for the efficient synthesis of NDP-uronic acids,and 11 NDP-uronic acids were successfully prepared in good yield and on a large scale.The prepared NDP-uronic acids can be used to explore new uronosyltransferases and synthesize uronic acids containing carbohydrates for fundamental research.
文摘Molecularly imprinted polymers(MIPs)exhibit high selectivity resulting from imprinted cavities and superior performance from functional materials,which have attracted much attention in many fields.However,the combination of MIPs film and functional materials is a great challenge.In this study,hemin/graphene hybrid nanosheets(H-GNs)were used to initiate the imprinted polymerization by catalyzing the generation of free radicals.Thus,MIPs using sulfamethoxazole as the template was directly prepared on the surface of H-GNs without any film modification.Most importantly,the template could be absorbed on the H-GNs to enhance the number of imprinted sites per unit surface area,which could improve the selectivity of MIPs film.Thus,the composites could exhibit high adsorption capacity(29.4 mg/g),imprinting factor(4.2)and excellent conductivity,which were modified on the surface of electrode for rapid,selective and sensitive detection of sulfamethoxazole in food and serum samples.The linear range was changed from 5μg/kg to 1 mg/g and the limit of detection was 1.2μg/kg.This sensor was free from interference caused by analogues of sulfamethoxazole,which provides a novel insight for the preparation of MIPs-based sensor and its application in food safety monitoring and human exposure study.
