Making peptide bonds is tightly controlled by genetic code and machinery which includes cofactors,ATP,and RNAs.In this regard,the stand-alone and genetic-code-independent peptide ligases constitute a new family of ren...Making peptide bonds is tightly controlled by genetic code and machinery which includes cofactors,ATP,and RNAs.In this regard,the stand-alone and genetic-code-independent peptide ligases constitute a new family of renegade peptide-bond makers.A prime example is butelase-1,an Asn/Asp(Asx)-specific ligase that structurally belongs to the asparaginyl endopeptidase family.Butelase-1 specifically recognizes a C-terminal Asx-containing tripeptide motif,Asn/Asp-Xaa-Yaa(Xaa and Yaa are any amino acids),to form a site-specific Asn-Xaa peptide bond either intramolecularly as cyclic proteins or intermolecularly as modified proteins.Our work in the past five years has validated that butelase-1 is a potent and versatile ligase.Here we review the advances in ligases,with a focus on butelase-1,and their applications in engineering bioactive peptides and precision protein modifications,antibody-drug conjugates,and live-cell labeling.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
采用生物信息学方法分析人锌指蛋白219(zinc finger protein 219,ZNF219)的空间结构、理化性质、结构域、蛋白质修饰位点以及蛋白质相互作用关系等信息。分析结果显示,ZNF219为一种碱性不稳定亲水蛋白,无信号肽,属于非跨膜蛋白,定位于...采用生物信息学方法分析人锌指蛋白219(zinc finger protein 219,ZNF219)的空间结构、理化性质、结构域、蛋白质修饰位点以及蛋白质相互作用关系等信息。分析结果显示,ZNF219为一种碱性不稳定亲水蛋白,无信号肽,属于非跨膜蛋白,定位于细胞核内,有9个zinc finger结构域,66个潜在的磷酸化位点,26个潜在的O-型糖基化位点,可能与HMGN1、MBD2等有相互作用。该研究通过多种生物信息学分析软件对ZNF219的结构及性质进行分析与预测,为研究其功能以及在癌症发生发展中的作用机制提供理论依据。展开更多
Over the past 20 years,great efforts have been invested in developing site-specific approaches to protein modification to dissect protein functions directly and accurately.Here,we report a proximitytriggered group tra...Over the past 20 years,great efforts have been invested in developing site-specific approaches to protein modification to dissect protein functions directly and accurately.Here,we report a proximitytriggered group transfer strategy from a sulfonium warhead to a Cysteine(Cys)residue of the target protein.With a guiding ligand,cargoes could be transferred selectively from a sulfonium center onto the Cys residue in the vicinity of their binding interface.The successful thalidomide transfer of sulfonium 1-X could be applied intracellularly for epidermal growth factor receptor degradation,highlighting the potential of group transfer strategy as a suite of chemical biology studies,including cell imaging,protein profiling,and protein degradation by simply employing different transferrable groups.展开更多
Site-selective modification of peptide/protein is a vital approach to disclose post-translational modifications(PTMs) and plays a crucial role in chemical biology, as well as drug development. Compared with synthetic ...Site-selective modification of peptide/protein is a vital approach to disclose post-translational modifications(PTMs) and plays a crucial role in chemical biology, as well as drug development. Compared with synthetic and chemical biology methods, chemical modification of native peptide/protein provides a more versatile approach to achieve late-stage diversification for functional studies. Lysine featured high nucleophilicity, frequency, and solvent accessibility, making its site-selective modification important but elusive. Herein, we reported a visible-light-driven and Cys-directed Lys site-selective stapling approach for peptide/protein. By cleavable Cys anchoring, site-selective Lys single-site modification was achieved, and this method could be applied to multi-functionalization.展开更多
Herein,we report a semi-synthetic strategy affording a nitrophorin 2(NP2)variant with a N,N'-bis(2-pyridylmethyl)amine(Dpa)ligand as sidechain selectively installed at position 27,which was assembled from a synthe...Herein,we report a semi-synthetic strategy affording a nitrophorin 2(NP2)variant with a N,N'-bis(2-pyridylmethyl)amine(Dpa)ligand as sidechain selectively installed at position 27,which was assembled from a synthetic peptide thioester bearing the Dpa ligand and an expressed protein segment via native chemical ligation.The semi-synthetic NP2 was able to accept the natural heme b cofactor and the Dpa ligand was able to bind Cu(Ⅱ)/Fe(Ⅲ)ions,leading to heteronuclear active site.展开更多
目的对小鼠精子发生相关蛋白3(spermatogenesis associated protein 3,Spata3)的序列进行分析,探讨其结构和功能。方法利用NCBI数据库比对小鼠Spata3的同源性、ExPASy ProtParam软件分析理化特征、SOPMA及GOR4预测空间构象、NetPhos3.1...目的对小鼠精子发生相关蛋白3(spermatogenesis associated protein 3,Spata3)的序列进行分析,探讨其结构和功能。方法利用NCBI数据库比对小鼠Spata3的同源性、ExPASy ProtParam软件分析理化特征、SOPMA及GOR4预测空间构象、NetPhos3.1及STRING数据库分析蛋白修饰位点及蛋白间相互作用关系等信息、免疫组化和免疫荧光染色检测其组织细胞定位。结果小鼠和人的Spata3基因CDS序列64%相同,是碱性不稳定亲水蛋白,无信号肽,属于非跨膜的胞内蛋白,主要定位于睾丸组织各级精子细胞核和胞质,以圆形精子细胞表达量最高。小鼠Spata3含1个内在无序区结构域,30个潜在的磷酸化位点,11个潜在的O-型糖基化位点,可能与Spata46、Spert等蛋白相互作用。结论小鼠Spata3是精子发生过程中的保守蛋白,可能调节精子发生变形过程。展开更多
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
基金supported by Academic Research Grant Tier 3(MOE2016-T3-1-003)from the Singapore Ministry of Education.
文摘Making peptide bonds is tightly controlled by genetic code and machinery which includes cofactors,ATP,and RNAs.In this regard,the stand-alone and genetic-code-independent peptide ligases constitute a new family of renegade peptide-bond makers.A prime example is butelase-1,an Asn/Asp(Asx)-specific ligase that structurally belongs to the asparaginyl endopeptidase family.Butelase-1 specifically recognizes a C-terminal Asx-containing tripeptide motif,Asn/Asp-Xaa-Yaa(Xaa and Yaa are any amino acids),to form a site-specific Asn-Xaa peptide bond either intramolecularly as cyclic proteins or intermolecularly as modified proteins.Our work in the past five years has validated that butelase-1 is a potent and versatile ligase.Here we review the advances in ligases,with a focus on butelase-1,and their applications in engineering bioactive peptides and precision protein modifications,antibody-drug conjugates,and live-cell labeling.
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.
文摘采用生物信息学方法分析人锌指蛋白219(zinc finger protein 219,ZNF219)的空间结构、理化性质、结构域、蛋白质修饰位点以及蛋白质相互作用关系等信息。分析结果显示,ZNF219为一种碱性不稳定亲水蛋白,无信号肽,属于非跨膜蛋白,定位于细胞核内,有9个zinc finger结构域,66个潜在的磷酸化位点,26个潜在的O-型糖基化位点,可能与HMGN1、MBD2等有相互作用。该研究通过多种生物信息学分析软件对ZNF219的结构及性质进行分析与预测,为研究其功能以及在癌症发生发展中的作用机制提供理论依据。
基金Weare grateful for the financial support from the Natural Science Foundation of China(grant nos.21778009,21977010,and 22007033)National Key Research and Development Program“Synthetic Biology”Key Special Project of China(grant no.2018YFA0902504)+3 种基金China Postdoctoral Science Foundation(grant no.2021M690220)the Natural Science Foundation of Guangdong Province(grant nos.2020A1515010522,2020A1515010766,2019A1515110487,and 2019A151-5111184)the Foundation for Basic and Applied Research of Guangdong Province(grant no.2019A1515110489)and the Shenzhen Science and Technology Innovation Committee(grant nos.JCYJ20180507181527112,JCYJ-201805081522131455,and JCYJ20170817172023838).
文摘Over the past 20 years,great efforts have been invested in developing site-specific approaches to protein modification to dissect protein functions directly and accurately.Here,we report a proximitytriggered group transfer strategy from a sulfonium warhead to a Cysteine(Cys)residue of the target protein.With a guiding ligand,cargoes could be transferred selectively from a sulfonium center onto the Cys residue in the vicinity of their binding interface.The successful thalidomide transfer of sulfonium 1-X could be applied intracellularly for epidermal growth factor receptor degradation,highlighting the potential of group transfer strategy as a suite of chemical biology studies,including cell imaging,protein profiling,and protein degradation by simply employing different transferrable groups.
基金supported by Guangdong Natural Science Funds for Distinguished Young Scholar (2018B030306017)the National Natural Science Foundation of China (22077144)+1 种基金Guangdong Provincial Key Laboratory of Chiral Molecule and Drug Discovery (2019B030301005)Key Research and Development Program of Guangdong Province (2020B1111110003)。
文摘Site-selective modification of peptide/protein is a vital approach to disclose post-translational modifications(PTMs) and plays a crucial role in chemical biology, as well as drug development. Compared with synthetic and chemical biology methods, chemical modification of native peptide/protein provides a more versatile approach to achieve late-stage diversification for functional studies. Lysine featured high nucleophilicity, frequency, and solvent accessibility, making its site-selective modification important but elusive. Herein, we reported a visible-light-driven and Cys-directed Lys site-selective stapling approach for peptide/protein. By cleavable Cys anchoring, site-selective Lys single-site modification was achieved, and this method could be applied to multi-functionalization.
基金financial support from the National Natural Science Foundation of China(Nos.22077036 and 22277029)is greatly acknowledged.
文摘Herein,we report a semi-synthetic strategy affording a nitrophorin 2(NP2)variant with a N,N'-bis(2-pyridylmethyl)amine(Dpa)ligand as sidechain selectively installed at position 27,which was assembled from a synthetic peptide thioester bearing the Dpa ligand and an expressed protein segment via native chemical ligation.The semi-synthetic NP2 was able to accept the natural heme b cofactor and the Dpa ligand was able to bind Cu(Ⅱ)/Fe(Ⅲ)ions,leading to heteronuclear active site.
文摘目的对小鼠精子发生相关蛋白3(spermatogenesis associated protein 3,Spata3)的序列进行分析,探讨其结构和功能。方法利用NCBI数据库比对小鼠Spata3的同源性、ExPASy ProtParam软件分析理化特征、SOPMA及GOR4预测空间构象、NetPhos3.1及STRING数据库分析蛋白修饰位点及蛋白间相互作用关系等信息、免疫组化和免疫荧光染色检测其组织细胞定位。结果小鼠和人的Spata3基因CDS序列64%相同,是碱性不稳定亲水蛋白,无信号肽,属于非跨膜的胞内蛋白,主要定位于睾丸组织各级精子细胞核和胞质,以圆形精子细胞表达量最高。小鼠Spata3含1个内在无序区结构域,30个潜在的磷酸化位点,11个潜在的O-型糖基化位点,可能与Spata46、Spert等蛋白相互作用。结论小鼠Spata3是精子发生过程中的保守蛋白,可能调节精子发生变形过程。
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.
