Simulating Multiple Substrate-Binding Events by γ-Glutamyltransferase Using Accelerated Molecular Dynamics.

Area of Science:

• Biochemistry
• Structural Biology
• Computational Biology

Background:

• γ-Glutamyltransferase (GGT) is a crucial enzyme with diverse physiological roles across species.
• While GGT crystal structures exist, the dynamic process of substrate binding to its active site remains uncharacterized.

Purpose of the Study:

• To investigate the molecular mechanisms of glutamine binding to *Escherichia coli* GGT.
• To elucidate the role of structural dynamics and specific interactions in GGT substrate recognition.

Main Methods:

• Accelerated molecular dynamics (aMD) simulations were employed to model glutamine binding to *E. coli* GGT.
• Analysis of binding events identified key structural motifs and conformational changes.

Main Results:

• Three distinct binding motifs involving polar residues were identified in the GGT active site, governing glutamine interaction.
• A lid-loop acts as a dynamic gate, controlling substrate access to the binding pocket, with partially open states accessible.
• The free energy cost for a fully open lid-loop conformation was estimated at 2.4 kcal/mol.

Conclusions:

• Substrate recognition by GGT is determined by a combination of specific electrostatic interactions and enzyme conformational dynamics.
• The lid-loop's gating mechanism plays a critical role in regulating substrate entry into the active site.
• This research provides novel insights into the dynamic aspects of enzyme-substrate interactions for GGT.