Exploring reaction pathways for O-GlcNAc transferase catalysis. A string method study.
The inverting O-GlcNAc glycosyltransferase (OGT) is an important post-translation enzyme, which catalyzes the transfer of N-acetylglucosamine from UDP-N-acetylglucosamine (UDP-GlcNAc) to the hydroxyl group of the Ser/Thr of cytoplasmic, nuclear, and mitochondrial proteins. In the past, three different catalytic bases were proposed for the reaction: His498, α-phosphate, and Asp554. In this study, we used hybrid quantum mechanics/molecular mechanics (QM/MM) Car-Parrinello molecular dynamics to investigate reaction paths using α-phosphate and Asp554 as the catalytic bases. The string method was used to calculate the free-energy reaction profiles of the tested mechanisms. During the investigations, an additional mechanism was observed. In this mechanism, a proton is transferred to α-phosphate via a water molecule. Our calculations show that the mechanism with α-phosphate acting as the base is favorable. This reaction has a rate-limiting free-energy barrier of 23.5 kcal/mol, whereas reactions utilizing Asp554 and water-assisted α-phosphate have barriers of 41.7 and 40.9 kcal/mol, respectively. Our simulations provide a new insight into the catalysis of OGT and may thus guide rational drug design of transition-state analogue inhibitors with potential therapeutic use.
†Central European Institute of Technology (CEITEC), Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic.|‡Faculty of Science - National Centre for Biomolecular Research, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic.
Manju
M
Kumari
†Central European Institute of Technology (CEITEC), Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic.|‡Faculty of Science - National Centre for Biomolecular Research, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic.
Stanislav
S
Kozmon
†Central European Institute of Technology (CEITEC), Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic.|‡Faculty of Science - National Centre for Biomolecular Research, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic.
Petr
P
Kulhánek
‡Faculty of Science - National Centre for Biomolecular Research, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic.
Jakub
J
Štepán
†Central European Institute of Technology (CEITEC), Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic.|§Institute of Chemistry, Slovak Academy of Sciences, Dúbravská cesta 9, 845 38 Bratislava, Slovak Republic.
Igor
I
Tvaroška
†Central European Institute of Technology (CEITEC), Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic.|‡Faculty of Science - National Centre for Biomolecular Research, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic.
Jaroslav
J
Koča
D002384:Catalysis; D006801:Humans; D056004:Molecular Dynamics Simulation; D017351:N-Acetylglucosaminyltransferases / Q000737:chemistry*; D011522:Protons; D011789:Quantum Theory; D014867:Water / Q000737:chemistry
D011522:Protons; D014867:Water; D017351:N-Acetylglucosaminyltransferases; C449880:O-GlcNAc transferase