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|Title:||A cascade enzymatic reaction scheme for irreversible transpeptidative protein ligation||Authors:||Xia, Yiyin
Tam, James P.
|Keywords:||Science::Biological sciences||Issue Date:||2023||Source:||Xia, Y., Li, F., Zhang, X., Balamkundu, S., Tang, F., Hu, S., Lescar, J., Tam, J. P. & Liu, C. (2023). A cascade enzymatic reaction scheme for irreversible transpeptidative protein ligation. Journal of the American Chemical Society, 145(12), 6838-6844. https://dx.doi.org/10.1021/jacs.2c13628||Project:||MOE2016-T3-1-003
|Journal:||Journal of the American Chemical Society||Abstract:||Enzymatic peptide ligation holds great promise in the study of protein functions and development of protein therapeutics. Owing to their high catalytic efficiency and a minimal tripeptide recognition motif, peptidyl asparaginyl ligases (PALs) are particularly useful tools for bioconjugation. However, as an inherent limitation of transpeptidases, PAL-mediated ligation is reversible, requiring a large excess of one of the ligation partners to shift the reaction equilibrium in the forward direction. Herein, we report a method to make PAL-mediated intermolecular ligation irreversible by coupling it to glutaminyl cyclase (QC)-catalyzed pyroglutamyl formation. In this method, the acyl donor substrate of PALs is designed to have glutamine at the P1' position of the Asn-P1'-P2' tripeptide PAL recognition motif. Upon ligation with an acyl acceptor substrate, the acyl donor substrate releases a leaving group in which the exposed N-terminal glutamine is cyclized by QC, quenching the Gln Nα-amine in a lactam. Using this method, PAL-mediated ligation can achieve near-quantitative yields even at an equal molar ratio between the two ligation partners. We have demonstrated this method for a wide range of applications, including protein-to-protein ligations. We anticipate that this cascade enzymatic reaction scheme will make PAL enzymes well suited for numerous new uses in biotechnology.||URI:||https://hdl.handle.net/10356/169065||ISSN:||0002-7863||DOI:||10.1021/jacs.2c13628||Schools:||School of Biological Sciences||Rights:||© 2023 American Chemical Society. All rights reserved.||Fulltext Permission:||none||Fulltext Availability:||No Fulltext|
|Appears in Collections:||SBS Journal Articles|
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