Oxygen diffusion pathways in a cofactor-independent dioxygenase.

Area of Science:

• Biochemistry and Molecular Biology
• Enzymology
• Protein Dynamics

Background:

• Molecular oxygen (O2) is crucial for numerous enzymatic reactions.
• Understanding O2 diffusion to enzyme active sites is key to enzyme function.
• Cofactor-independent oxygenases, like DpgC, are rare and mechanistically intriguing.

Purpose of the Study:

• To elucidate the O2 diffusion pathways to the active site of DpgC.
• To investigate the role of enzyme structure in O2 transport and substrate activation.
• To compare O2 access mechanisms in DpgC with other O2-utilizing proteins.

Main Methods:

• Extensive molecular dynamics (MD) simulations.
• Site-directed mutagenesis experiments.
• Xenon-binding data analysis.
• Kinetic measurements.

Main Results:

• Identified three primary pathways and four access points for O2 entry into DpgC.
• MD simulations revealed dynamic hydrophobic pockets guiding O2 to the substrate.
• Simulated pathway usage correlated with experimental kinetic data from mutants.

Conclusions:

• DpgC utilizes a network of dynamic channels for O2 delivery to its active site.
• The O2 access mechanism in DpgC shares common themes with other oxygenases, emphasizing conserved transport strategies.
• Enzyme architecture dictates regio- and stereoselectivity in O2 activation, even without cofactors or metals.