Dissecting the interaction between transglutaminase 2 and fibronectin

Area of Science:

• Biochemistry
• Molecular Biology
• Cell Biology

Background:

• Transglutaminase 2 (TG2) is an extracellular enzyme mediating cell-matrix interactions.
• TG2 binds to fibronectin (FN), particularly its 45 kDa gelatin-binding domain (45FN).
• The precise location of the FN-binding site on TG2 has been previously debated.

Purpose of the Study:

• To map the fibronectin (FN) interaction site on human transglutaminase 2 (TG2).
• To identify specific amino acid residues critical for the high-affinity binding of TG2 to FN.
• To elucidate the structural basis of the TG2-FN interaction.

Main Methods:

• Hydrogen/deuterium exchange mass spectrometry (HDX-MS) to map the FN interaction site.
• Site-directed mutagenesis to alter specific TG2 residues.
• Surface plasmon resonance (SPR) analysis to quantify binding affinity.

Main Results:

• Confirmed the FN-binding site is in the N-terminal domain of TG2.
• Identified TG2 residues K30, R116, and H134 as essential for high-affinity FN binding.
• Simultaneous mutation of K30, R116, and H134 decreased 45FN binding by over 2000-fold.
• Identified additional contributing residues in the catalytic core domain, expanding the TG2-FN binding interface.

Conclusions:

• Precisely mapped the FN interaction site on human TG2.
• Identified key residues (K30, R116, H134) and extended the binding interface for TG2-FN interaction.
• Provides novel insights into the molecular mechanisms governing TG2-FN binding and cell-matrix interactions.