Exploring reaction pathways for O-GlcNAc transferase catalysis. A string method study

Area of Science:

• Biochemistry
• Enzymology
• Computational Chemistry

Background:

• O-GlcNAc glycosyltransferase (OGT) is a crucial post-translational enzyme.
• OGT catalyzes the transfer of N-acetylglucosamine to Ser/Thr residues in proteins.
• Previous studies proposed His498, α-phosphate, or Asp554 as catalytic bases for OGT.

Purpose of the Study:

• To investigate the catalytic mechanisms of OGT using computational methods.
• To determine the most favorable reaction pathway for OGT-catalyzed glycosylation.
• To provide insights for the rational design of OGT inhibitors.

Main Methods:

• Hybrid quantum mechanics/molecular mechanics (QM/MM) Car-Parrinello molecular dynamics simulations.
• String method to calculate free-energy reaction profiles.
• Evaluation of reaction mechanisms involving α-phosphate and Asp554 as catalytic bases.

Main Results:

• Identified a novel mechanism involving water-assisted proton transfer to α-phosphate.
• Calculations revealed that α-phosphate acting as the catalytic base is the most favorable mechanism.
• The rate-limiting free-energy barrier for the α-phosphate mechanism is 23.5 kcal/mol, significantly lower than other proposed mechanisms.

Conclusions:

• The α-phosphate mechanism is the predominant catalytic pathway for OGT.
• Computational simulations offer valuable insights into OGT's enzymatic activity.
• Findings can guide the development of transition-state analogue inhibitors for therapeutic applications.