TY  - JOUR
TI  - In vitro Crosslinking Reactions and Substrate Incorporation Assays for The Identification of Transglutaminase-2 Protein Substrates
AU  - Willis, William  L.
AU  - Foster, Abigail 
AU  - Henry, Caitlin 
AU  - Wu, Lai Chu 
AU  - Jarjour, Wael 
VL  - 10
IS  - 12
PY  - 2020
DA  - 2020/06/20
SP  - e3657
C1  - Bio-protocol 2020;10:e3657
DO  - 10.21769/BioProtoc.3657
UR  - https://doi.org/10.21769/BioProtoc.3657
AB  - Transglutaminase (TG2) catalyzes protein crosslinking between glutamyl and lysyl residues. Catalytic activity occurs via a transamidation mechanism resulting in the formation of isopeptide bonds. Since TG2-mediated transamidation is of mechanistic importance for a number of biological processes, assays that enable rapid and efficient identification and characterization of candidate substrates are an important first-step to uncovering the function of crosslinked proteins. Herein we describe an optimized and flexible protocol for in vitro TG2 crosslink reactions and substrate incorporation assays. We have previously employed these techniques in the identification of the protein high mobility group box 1 (HMGB1) as a TG2 substrate. However, the protocol can be adapted for identification of any candidate transamidation substrate.
KW  - Transglutaminase
KW  - Transamidation
KW  - Post-translational modification
KW  - High mobility group box 1
KW  - Isopeptide
KW  - In vitro protein crosslinking
JF  - Bio-protocol
SN  - 2331-8325
PB  - Bio-protocol LLC.
BIO101  - False