Histidine(His)bears a uniquely electron-deficient imidazole side chain and plays essential roles in protein interactions and enzyme-catalyzed processes.Modification of His C_(2)position offers a useful method to fine-...Histidine(His)bears a uniquely electron-deficient imidazole side chain and plays essential roles in protein interactions and enzyme-catalyzed processes.Modification of His C_(2)position offers a useful method to fine-tune histidine residues of proteins for their structural and functional study.Due to the moderately nucleophilic imidazole group,the chemoselective modification of histidine in proteins remains particularly challenging.Herein,we report a highly efficient method for the semisynthesis of chitin-binding protein21(CBP21)bearing various groups at the C_(2)position of His28.A combination of modern radical-mediated C-H alkylation and recombinant protein engineering offers a powerful strategy to decipher His functions.展开更多
Expressed protein ligation(EPL)provides a powerful tool to access large-size proteins with precise structures.Existing methods for constructing the critical protein thioester for EPL have predominantly relied on the r...Expressed protein ligation(EPL)provides a powerful tool to access large-size proteins with precise structures.Existing methods for constructing the critical protein thioester for EPL have predominantly relied on the recombinant intein fusion expressed in Escherichia coli(E.coli).Despite its powerful applications,the expression of thioester derived from eukaryotic protein in E.coli inherently suffers from its limited solubility,the inactivity of intein,premature hydrolysis and low yields.To overcome these obstacles,we present herein the facile one-flask synthesis of inaccessible proteinα-thioester via a SUMO-protein-intein(SPI)sandwich model.The utility of SUMO enhances the protein fusion yield and solubility,prevents premature hydrolysis and simplifies the purification process.The inaccessible protein thioester with internal Cys residues can be readily produced and is compatible with the EPL-desulfurization protocol used to prepare complex proteins,which is otherwise difficult to obtain using traditional methods.Its utility has been highlighted through the synthesis of human granulocyte colony-stimulating factor(G-CSF).展开更多
Naturally occurring interleukin-2(IL-2)is a pleiotropic glycoprotein that regulates immune responses by controlling the differentia-tion and homeostasis of T cells.Non-glycosylated IL-2 has been used in clinical setti...Naturally occurring interleukin-2(IL-2)is a pleiotropic glycoprotein that regulates immune responses by controlling the differentia-tion and homeostasis of T cells.Non-glycosylated IL-2 has been used in clinical settings for three decades.However,the function of the O-glycan of native IL-2 remains elusive.Herein,to stress this issue,we report a highly efficient semi-synthesis of homogeneous glycosylated IL-2 with various glycoproteoforms on a multi-milligram scale.The glycopeptide fragment was prepared by chemical synthesis and then merged with recombinant fragment via a serine ligation to generate the desired glycoprotein in a single opera-tion.Biological evaluation of the homogenous glycoprotein library reveals that the activity of IL-2 in activating individual T cell subset is glycan dependent,thus highlighting the possibility of further improving current clinical medicine.展开更多
Escherichia coli(E. coli) DH5α has been recognized as a non-pathogenic bacterial strain with tumor colonization ability. However, whether such a bacteria-driven drug-delivery system can improve the targeting of tumor...Escherichia coli(E. coli) DH5α has been recognized as a non-pathogenic bacterial strain with tumor colonization ability. However, whether such a bacteria-driven drug-delivery system can improve the targeting of tumor therapy or not remains essentially untouched. Herein, a series of zinc phthalocyanine(ZnPc) photosensitizers with different numbers of charges were prepared and their electrostatic adhesion properties on E. coli were investigated via measuring their fluorescence intensities by flow cytometer. Among these ZnPc photosensitizers investigated, the ZnPc conjugate with four positive charges(named ZnPc-IR710) exhibited the highest loading capacity and the best fluorescence imaging performance of E. coli. With the help of E. coli, E. coli@ZnPcIR710 presented a significantly enhanced cytotoxicity on human breast cancer MCF-7 cells compared with ZnPc-IR710(survival rate of tumor cells was 39% vs. 57% at a concentration of 50 nmol L-1). Moreover, in vivo study showed that E. coli@ZnPc-IR710 remarkably inhibited the tumor growth and resulted in a complete tumor growth suppress in subcutaneous mouse 4T1 breast tumor model. These results demonstrated the great promise of bacterial-guided photodynamic therapy(PDT) in the treatment of solid tumors, and provide a unique strategy to enhance the antitumor efficacy of PDT by utilizing bacterial vectors in tumors.展开更多
基金supported by the National Science Fund for National Natural Science Foundation of China(grant nos.92253302,22225701,22077080 and 22477078)the Shanghai Pilot Program for Basic Research-Shanghai Jiao Tong University(grant no.21TQ1400210)+1 种基金the Interdisciplinary Program of Shanghai Jiao Tong University(grant no.20230102)the Shanghai Municipal Science and Technology Major Project and the Department of Human Resources,and Social Security of Sichuan Province,China.
文摘Histidine(His)bears a uniquely electron-deficient imidazole side chain and plays essential roles in protein interactions and enzyme-catalyzed processes.Modification of His C_(2)position offers a useful method to fine-tune histidine residues of proteins for their structural and functional study.Due to the moderately nucleophilic imidazole group,the chemoselective modification of histidine in proteins remains particularly challenging.Herein,we report a highly efficient method for the semisynthesis of chitin-binding protein21(CBP21)bearing various groups at the C_(2)position of His28.A combination of modern radical-mediated C-H alkylation and recombinant protein engineering offers a powerful strategy to decipher His functions.
基金supported by the National Science Fund for Distinguished Young Scholars(22225701)the Interdisciplinary Program of Shanghai Jiao Tong University(20230102)+3 种基金the National Natural Science Foundation of China(22077080,92253302 and 22275122)the Shanghai Pilot Program for Basic Research Shanghai Jiao Tong University(21TQ1400210)the Special Projects of the Central Government in Guidance of Local Science and Technology Development(2021Szvup077)the China Post-doctoral Science Foundation(2023M732214).
文摘Expressed protein ligation(EPL)provides a powerful tool to access large-size proteins with precise structures.Existing methods for constructing the critical protein thioester for EPL have predominantly relied on the recombinant intein fusion expressed in Escherichia coli(E.coli).Despite its powerful applications,the expression of thioester derived from eukaryotic protein in E.coli inherently suffers from its limited solubility,the inactivity of intein,premature hydrolysis and low yields.To overcome these obstacles,we present herein the facile one-flask synthesis of inaccessible proteinα-thioester via a SUMO-protein-intein(SPI)sandwich model.The utility of SUMO enhances the protein fusion yield and solubility,prevents premature hydrolysis and simplifies the purification process.The inaccessible protein thioester with internal Cys residues can be readily produced and is compatible with the EPL-desulfurization protocol used to prepare complex proteins,which is otherwise difficult to obtain using traditional methods.Its utility has been highlighted through the synthesis of human granulocyte colony-stimulating factor(G-CSF).
基金supported by the National Natural Science Foundation of China(22077080,21907064 and 22107068)Special Projects of the Central Government in Guidance of Local Science and Technology Development(2021Szvup077).
文摘Naturally occurring interleukin-2(IL-2)is a pleiotropic glycoprotein that regulates immune responses by controlling the differentia-tion and homeostasis of T cells.Non-glycosylated IL-2 has been used in clinical settings for three decades.However,the function of the O-glycan of native IL-2 remains elusive.Herein,to stress this issue,we report a highly efficient semi-synthesis of homogeneous glycosylated IL-2 with various glycoproteoforms on a multi-milligram scale.The glycopeptide fragment was prepared by chemical synthesis and then merged with recombinant fragment via a serine ligation to generate the desired glycoprotein in a single opera-tion.Biological evaluation of the homogenous glycoprotein library reveals that the activity of IL-2 in activating individual T cell subset is glycan dependent,thus highlighting the possibility of further improving current clinical medicine.
基金supported by the National Natural Science Foundation of China (81572944, 21471033, 21877113 and 81971983)the CAS/SAFEA International Partnership Program for Creative Research Teams, the High-Level Entrepreneurship and Innovation Talents Projects in Fujian Province (2018-8-1)the FJIRSM&IUE Joint Research Fund (RHZX-2018-004)。
文摘Escherichia coli(E. coli) DH5α has been recognized as a non-pathogenic bacterial strain with tumor colonization ability. However, whether such a bacteria-driven drug-delivery system can improve the targeting of tumor therapy or not remains essentially untouched. Herein, a series of zinc phthalocyanine(ZnPc) photosensitizers with different numbers of charges were prepared and their electrostatic adhesion properties on E. coli were investigated via measuring their fluorescence intensities by flow cytometer. Among these ZnPc photosensitizers investigated, the ZnPc conjugate with four positive charges(named ZnPc-IR710) exhibited the highest loading capacity and the best fluorescence imaging performance of E. coli. With the help of E. coli, E. coli@ZnPcIR710 presented a significantly enhanced cytotoxicity on human breast cancer MCF-7 cells compared with ZnPc-IR710(survival rate of tumor cells was 39% vs. 57% at a concentration of 50 nmol L-1). Moreover, in vivo study showed that E. coli@ZnPc-IR710 remarkably inhibited the tumor growth and resulted in a complete tumor growth suppress in subcutaneous mouse 4T1 breast tumor model. These results demonstrated the great promise of bacterial-guided photodynamic therapy(PDT) in the treatment of solid tumors, and provide a unique strategy to enhance the antitumor efficacy of PDT by utilizing bacterial vectors in tumors.
