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  • Review Article
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Nature-inspired protein ligation and its applications

Nature Reviews Chemistry volume 7, pages 234–255 (2023)Cite this article

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Abstract

The ability to manipulate the chemical composition of proteins and peptides has been central to the development of improved polypeptide-based therapeutics and has enabled researchers to address fundamental biological questions that would otherwise be out of reach. Protein ligation, in which two or more polypeptides are covalently linked, is a powerful strategy for generating semisynthetic products and for controlling polypeptide topology. However, specialized tools are required to efficiently forge a peptide bond in a chemoselective manner with fast kinetics and high yield. Fortunately, nature has addressed this challenge by evolving enzymatic mechanisms that can join polypeptides using a diverse set of chemical reactions. Here, we summarize how such nature-inspired protein ligation strategies have been repurposed as chemical biology tools that afford enhanced control over polypeptide composition.

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Fig. 1: An overview of the enzymatic toolbox for protein ligation-mediated polypeptide engineering covered in this Review.
Fig. 2: Biochemistry of protein and peptide ligations mediated by sortase and butelase-1.
Fig. 3: Biochemistry of protease-based protein and peptide ligations.
Fig. 4: Biochemistry of the isopeptide bond formation mediated by transglutaminases.
Fig. 5: Molecular superglue-mediated isopeptide bond formation.
Fig. 6: Ubiquitin and ubiquitin-like modifier ligase-mediated isopeptide bond formation and its applications.
Fig. 7: Mechanism of intein-mediated protein ligation.
Fig. 8: Split intein-based strategies for generating semisynthetic proteins.
Fig. 9: Strategies for conditional protein splicing (CPS).
Fig. 10: Cellular protein manipulation strategies using inteins.

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Acknowledgements

The authors thank M. Luo for critical reading of the manuscript. Work in the David Laboratory is supported by the Josie Robertson Foundation, the Pershing Square Sohn Cancer Research Alliance, the NIH (CCSG core grant P30 CA008748, MSK SPORE P50 CA192937, R21 DA044767 and R35 GM138386), the Parker Institute for Cancer Immunotherapy and the Anna Fuller Trust. In addition, the David Laboratory is supported by the William H. Goodwin and Alice Goodwin Commonwealth Foundation for Cancer Research and by the Center for Experimental Therapeutics at MSKCC. R.P. is supported by the Novo Nordisk Foundation grant NNF20OC0061064. The Zheng laboratory is supported by OSUCCC startup funds.

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Authors and Affiliations

  1. Chemical Biology Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA

    Rasmus Pihl & Yael David

  2. Department of Biomedicine, Aarhus University, Aarhus C, Denmark

    Rasmus Pihl

  3. Department of Radiation Oncology, College of Medicine, The Ohio State University, Columbus, OH, USA

    Qingfei Zheng

  4. Center for Cancer Metabolism, James Comprehensive Cancer Center, The Ohio State University, Columbus, OH, USA

    Qingfei Zheng

  5. Department of Biological Chemistry and Pharmacology, College of Medicine, The Ohio State University, Columbus, OH, USA

    Qingfei Zheng

  6. Department of Pharmacology, Weill Cornell Medicine, New York, NY, USA

    Yael David

  7. Department of Physiology, Biophysics and Systems Biology, Weill Cornell Medicine, New York, NY, USA

    Yael David

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Pihl, R., Zheng, Q. & David, Y. Nature-inspired protein ligation and its applications. Nat Rev Chem 7, 234–255 (2023). https://doi.org/10.1038/s41570-023-00468-z

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