SNAC-tag for Sequence-specific Chemical Protein Cleavage

By Bobo Dang, M. Mravic, Hailin Hu, N. Schmidt, Bruk Mensa, William F. DeGrado. 2019

70 citations recorded by the source corpus.

Site-specific protein cleavage is essential for many protein-production protocols and typically requires proteases. We report the development of a chemical protein-cleavage method that is achieved through the use of a sequence-specific nickel-assisted cleavage (SNAC)-tag. We demonstrate that the SNAC-tag can be inserted before both water-soluble and membrane proteins to achieve fusion protein cleavage under biocompatible conditions with efficiency comparable to that of enzymes, and that the method works even when enzymatic cleavages fail. SNAC-tags allow for versatile sequence-specific cleavage of soluble and membrane proteins with Ni2+ under biocompatible conditions, bypassing enzymatic cleavage and enabling cleavage in situations where commonly used enzymes fail.

Subfields: Molecular Biology, Radiology, Nuclear Medicine and Imaging, Computational Biology & Bioinformatics

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In Molecular Biology

1. Minkyung Baek
2. J. Dauparas
3. A. Kang
4. Frank DiMaio
5. L. Milles
6. Lauren P. Carter
7. B. Wicky
8. A. Courbet

In Computational Biology & Bioinformatics

1. A. Kang
2. Frank DiMaio
3. J. Dauparas
4. L. Milles
5. Lauren P. Carter
6. Minkyung Baek
7. B. Wicky
8. David Baker

Citations in the corpus

• Robust deep learning based protein sequence design using ProteinMPNN (2022, neutral): “6xhis affinity tags were removed via on-bead SNAC (26) cleavage.”
• Hallucinating protein assemblies (2022, methodology): “All subcloning reactions resulted in C-terminally HIS-tagged constructs, either as: MSG-design-GSHHHHHH (entry vector LM670) or a MSG-design-GSGSHHWGSTHHHHHH (entry vector LM627), where the underlined sequence is the SNAC-tag used for cleaving the HIS-tag for crystallization ( 36 ).”
• Reconfigurable asymmetric protein assemblies through implicit negative design (2021, unknown): “)HHHHHH] were cleaved chemically by on-bead nickelassisted cleavage (44): Nickel-bound designs were washed with 10 column volumes of lysis buffer followed by 5 column volumes of 20mM Tris pH 8.”