双酶修饰制备纳米抗体靶向成像蛋白簇

doi:10.3969/j.issn.0253-2778.2019.04.006

中国科学技术大学学报 ›› 2019, Vol. 49 ›› Issue (4): 297-301.DOI: 10.3969/j.issn.0253-2778.2019.04.006

双酶修饰制备纳米抗体靶向成像蛋白簇

田瑶，武田田，刘扬中

中国科学技术大学化学与材料科学学院，安徽合肥 230026

收稿日期:2018-04-12 修回日期:2018-09-14 接受日期:2018-09-14 出版日期:2019-04-30 发布日期:2018-09-14
通讯作者: 刘扬中
作者简介:田瑶，男，1993年生，硕士.研究方向：生物无机化学.E-mail：tianyao@mail.ustc.edu.cn
基金资助:
国家自然科学基金（21573213），大科学装置联合基金(U1332210)资助.

Nanobody cluster imaging probe based on sortase and transglutaminase

TIAN Yao, WU Tiantian, LIU Yangzhong

School of Chemistry and Materials Science, University of Science and Technology of China, Hefei 230026, China

Received:2018-04-12 Revised:2018-09-14 Accepted:2018-09-14 Online:2019-04-30 Published:2018-09-14

摘要/Abstract

摘要： 通过分选酶和谷氨酰胺转氨酶催化反应制备了一种由纳米抗体和绿色荧光蛋白sfGFP组成的靶向成像探针.通过设计一类具有两种酶识别结构的树枝状聚氨基酸底物来制备偶联了纳米抗体和sfGFP的蛋白簇.该靶向蛋白簇能够特异性结合靶细胞并呈现绿色荧光信号.在肿瘤异种移植的小鼠模型中，该靶向蛋白簇能够快速富集到肿瘤部位，并在9 h内逐渐被代谢排出.

关键词: 分选酶, 谷氨酰胺转氨酶, 纳米抗体, 定点修饰, 靶向成像

Abstract: Sortase and transglutaminase was rendered for preparing a targeted protein cluster probe, which is composed nanobody and green fluorescence protein (sfGFP). A unique substrate, which can be recognized by both sortase and transglutaminase, was designed for nanobody and sfGFP conjunction by solid phase synthesis. The targeted protein cluster probe, namely KK-sfGFP(Nb), can specificly bind to the EGFR overexpressed cells and shows green fluorescence as signal. The KK-sfGFP(Nb) exhibits fast accumulation in mouse xenograft tumor and the fluorescence signal gradually metabolizes within 9 h.

Key words: sortase, transglutaminase, nanobody, site-specific modification, target imaging

中图分类号:

R979.1

田瑶，武田田，刘扬中. 双酶修饰制备纳米抗体靶向成像蛋白簇[J]. 中国科学技术大学学报, 2019, 49(4): 297-301.

TIAN Yao, WU Tiantian, LIU Yangzhong. Nanobody cluster imaging probe based on sortase and transglutaminase[J]. Journal of University of Science and Technology of China, 2019, 49(4): 297-301.

参考文献

［1］
VERONESE F M. Peptide and protein PEGylation: A review of problems and solutions[J]. Biomaterials, 2001, 22 (5): 405-417.
[2] FILPULA D. Antibody engineering and modification technologies[J]. Biomolecular Engineering, 2007, 24 (2): 201-215.
[3] SCHMIDT M, TOPLAK A, QUAEDFLIEG P J L M, et al. Enzyme-mediated ligation technologies for peptides and proteins[J]. Current Opinion in Chemical Biology, 2017, 38: 1-7.
[4] GLASGOW J E, SALIT M L, COCHRAN J R. In vivo site-specific protein tagging with diverse amines using an engineered sortase variant[J]. Journal of the American Chemical Society, 2016, 138 (24): 7496-7499.
[5] MAO H Y, HART S A, SCHINK A, et al. Sortase-mediated protein ligation: A new method for protein engineering [J]. Journal of the American Chemical Society, 2004, 126 (9): 2670-2671.
[6] THERIEN A, BEDARD M, CARIGNAN D, et al. A versatile papaya mosaic virus (PapMV) vaccine platform based on sortase-mediated antigen coupling[J]. Journal of Nanobiotechnology, 2017, 15:54.
[7] CHEN Q, SUN Q, MOLINO N M, et al. Sortase A-mediated multi-functionalization of protein nanoparticles [J]. Chemical Communications, 2015, 51 (60): 12107-12110.
[8] TAKI M, SHIOTA M, TAIRA K. Transglutaminase-mediated N- and C-terminal fluorescein labeling of a protein can support the native activity of the modified protein[J]. Protein Engineering Design & Selection, 2004, 17 (2): 119-126.
[9] WU T T, HUANG H, SHENG Y, et al. Transglutaminase mediated PEGylation of nanobodies for targeted nano-drug delivery[J]. Journal of Materials Chemistry B, 2018, 6 (7): 1011-1017.
[10] JEGER S, ZIMMERMANN K, BLANC A, et al. Site-specific and stoichiometric modification of antibodies by bacterial transglutaminase[J]. Angewandte Chemie International Edition, 2010, 49 (51): 9995-9997.
[11] DENNLER P, CHIOTELLIS A, FISCHER E, et al. Transglutaminase-based chemo-enzymatic conjugation approach yields homogeneous antibody-drug conjugates[J]. Bioconjugate Chemistry, 2014, 25 (3): 569-578.
[12] HAMERS-CASTERMAN C, ATARHOUCH T, MUYLDERMANS S, et al. Naturally occurring antibodies devoid of light chains[J]. Nature, 1993, 363 (6428): 446-448.
[13] VAN AUDENHOVE I, GETTEMANS J. Nanobodies as versatile tools to understand, diagnose, visualize and treat cancer[J]. Ebiomedicine, 2016, 8: 40-48.
[14] AL-HOMSI L, AL-ASSAD J M, KWEIDER M, et al. Construction of pRSET-sfGFP plasmid for fusion-protein expression[J]. Jordan Journal of Biological Sciences, 2012, 5: 279-288.
[15] WITTE M D, WU T F, GUIMARAES C P, et al. Site-specific protein modification using immobilized sortase in batch and continuous-flow systems[J]. Nature Protocols, 2015, 10 (3):508-516.
[16] MERRIFIELD B. Solid phase synthesis [J]. Science, 1985, 5 (5): 353-376.
[17] ROTHBAUER U, ZOLGHADR K, TILLIB S, et al. Targeting and tracing antigens in live cells with fluorescent nanobodies[J]. Nature Methods, 2006, 3 (11): 887-889.

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双酶修饰制备纳米抗体靶向成像蛋白簇

Nanobody cluster imaging probe based on sortase and transglutaminase

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