Academic Journal
MccB-catalyzed C-terminal Thioesterification for Protein Bioconjugation.
| Τίτλος: | MccB-catalyzed C-terminal Thioesterification for Protein Bioconjugation. |
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| Συγγραφείς: | Yang D; Department of Biochemistry, University of Wisconsin-Madison, Madison, WI, USA., Weeks AM; Department of Biochemistry, University of Wisconsin-Madison, Madison, WI, USA. |
| Πηγή: | Current protocols [Curr Protoc] 2026 Jun; Vol. 6 (6), pp. e70398. |
| Τύπος έκδοσης: | Journal Article |
| Γλώσσα: | English |
| Στοιχεία περιοδικού: | Publisher: John Wiley & Sons Country of Publication: United States NLM ID: 101773894 Publication Model: Print Cited Medium: Internet ISSN: 2691-1299 (Electronic) Linking ISSN: 26911299 NLM ISO Abbreviation: Curr Protoc Subsets: MEDLINE |
| Imprint Name(s): | Original Publication: Hoboken, NJ : John Wiley & Sons, [2021]- |
| Ιατρικοί όροι (MeSH): | Esters*/chemistry , Esters*/metabolism , Proteins*/chemistry , Proteins*/metabolism , Sulfhydryl Compounds*/chemistry , Sulfhydryl Compounds*/metabolism, Esterification |
| Περίληψη: | Protein bioconjugation enables the installation of functional groups, probes, and payloads that cannot be genetically encoded, thereby enabling the study and manipulation of biological systems. An important class of intermediates in protein bioconjugation is C-terminal protein α-thioesters, which are kinetically stable yet thermodynamically activated as electrophiles for C-terminal functionalization. An ATP-driven, MccB-catalyzed enzymatic system for synthesis of protein C-terminal thioesters was recently developed to complement existing tools for chemical and intein-mediated synthesis of C-terminal thioesters. This system relies on modification of the target protein with a thioesterification C-terminal handle (TeCH tag) that is specifically recognized by MccB, enabling conversion of the C-terminal α-carboxylate to an α-thioester. Thioesters generated with the MccB/TeCH tag system can be integrated with well-established bioconjugation methods, including expressed protein ligation and enzyme-catalyzed expressed protein ligation, expanding the scope and versatility of this approach. This article outlines the steps for applying the MccB/TeCH tag system for epitope-specific C-terminal thioesterification and for the use of MccB-generated thioesters for expressed protein ligation and enzyme-catalyzed expressed protein ligation. © 2026 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: MccB-catalyzed protein C-terminal thioesterification Alternate Protocol 1: MccB-catalyzed expressed protein ligation Alternate Protocol 2: Stepwise MccB-catalyzed protein C-terminal thioesterification followed by expressed protein ligation Support Protocol 1: Expression and purification of MccB Basic Protocol 2: MccB- and subtiligase-catalyzed expressed protein ligation. (© 2026 The Author(s). Current Protocols published by Wiley Periodicals LLC.) |
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| Grant Information: | NIH; David and Lucille Packard Foundation; Ono Pharma Foundation |
| Contributed Indexing: | Keywords: expressed protein ligation; protein C terminus; protein bioconjugation; protein thioester; subtiligase |
| Substance Nomenclature: | 0 (Esters) 0 (Proteins) 0 (Sulfhydryl Compounds) |
| Entry Date(s): | Date Created: 20260611 Date Completed: 20260612 Latest Revision: 20260726 |
| Update Code: | 20260726 |
| PubMed Central ID: | PMC13255007 |
| DOI: | 10.1002/cpz1.70398 |
| PMID: | 42273971 |
| Βάση Δεδομένων: | MEDLINE |
| ISSN: | 2691-1299 |
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| DOI: | 10.1002/cpz1.70398 |