ISSN 0253-2778

CN 34-1054/N

Open AccessOpen Access JUSTC Original Paper

Nanobody cluster imaging probe based on sortase and transglutaminase

Cite this:
https://doi.org/10.3969/j.issn.0253-2778.2019.04.006
  • Received Date: 12 April 2018
  • Accepted Date: 14 September 2018
  • Rev Recd Date: 14 September 2018
  • Publish Date: 30 April 2019
  • 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.
    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.
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  • [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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Catalog

    [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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