Mechanism of Proton Release during Water Oxidation in Photosystem II

Academic Background

Photosystem II (PSII) is the only enzyme in nature capable of catalyzing water splitting, a reaction that not only releases oxygen but also provides electrons for biomass synthesis. The water-splitting reaction releases protons into the thylakoid lumen, forming a proton-motive force (PMF) that drives ATP synthesis. Despite significant advances in the structural and functional studies of PSII, the mechanism of key steps in water oxidation, particularly the deprotonation process, remains controversial. This study combines quantum/classical (QM/MM) free energy calculations and atomic molecular dynamics (MD) simulations to reveal how the oxygen-evolving manganese-calcium cluster (Mn4O5Ca) in PSII transports protons to the thylakoid lumen via conserved carboxylates and water arrays. The study also identifies local proton storage sites and molecular gating mechanisms that prevent wasteful proton backflow.

Source of the Paper

This paper was co-authored by Friederike Allgöwer, Maximilian C. Pöverlein, A. William Rutherford, and Ville R. I. Kaila, from the Department of Biochemistry and Biophysics at Stockholm University and the Department of Life Sciences at Imperial College London. The paper was published on December 19, 2024, in the Proceedings of the National Academy of Sciences (PNAS) under the title Mechanism of Proton Release during Water Oxidation in Photosystem II.

Research Process

1. Research Design

This study aims to reveal the mechanism of proton release during water oxidation in PSII through multiscale simulation methods. The research combines classical molecular dynamics (MD) simulations with quantum/classical (QM/MM) free energy calculations, focusing on the proton transfer pathways in the Mn4O5Ca cluster of PSII.

2. Molecular Dynamics Simulations

The study first conducted microsecond-scale MD simulations of PSII embedded in a lipid membrane surrounded by water molecules and ions. The simulation system consisted of approximately 535,000 atoms, simulating PSII structures in different oxidation states (S2, S3, S4) and protonation states (Asp61D1, Glu312D2, Glu65D1, Glu310D2, Asp224PsbO). Each simulation lasted 200 nanoseconds, totaling about 10 microseconds.

3. Quantum/Classical Simulations

Based on the MD simulations, the study performed QM/MM free energy calculations and ab initio molecular dynamics (QM/MM-MD) simulations. The QM/MM model included approximately 16,500 classical atoms and 258 to 276 quantum atoms, simulating the proton transfer pathway in the CL1 channel of PSII. The QM region included proton acceptors (Asp61D1, Glu312D2, Glu65D1, Glu310D2, Asp224PsbO) and their coordinating residues.

4. Free Energy Calculations

The study used umbrella sampling to calculate the free energy profiles of the proton transfer pathways. The reaction coordinate was defined as a linear combination of all bond formation/breaking distances along the proton transfer pathway. The free energy profiles were calculated using the weighted histogram analysis method (WHAM).

Key Results

1. Proton Transfer Pathway

The study revealed that the CL1 channel in PSII supports water-mediated proton release via a chain of carboxylates (Asp61D1, Glu312D2, Glu65D1, Glu310D2/Asp224PsbO). The oxidation of Tyr161D1 (YZ) lowers the kinetic barrier for proton transfer, driven by electric field effects.

2. Molecular Gating Mechanism

Glu65D1 was identified as a local molecular gate that controls proton transfer to the thylakoid lumen. Protonation of Glu65D1 induces a conformational change, causing its side chain to swing toward Glu310D2, thereby establishing proton connectivity to the lumen. This conformational change prevents proton backflow from the lumen to the Mn4O5Ca cluster.

3. Electric Field Effects

The study found that the electric field triggered by YZ oxidation enhances proton transfer in the CL1 channel. The electric field strength ranges from 1 to 2 V Å⁻¹, with YZ oxidation enhancing the field effect by approximately 0.5 V Å⁻¹. The electric field effect modulates the proton transfer barrier by inducing conformational changes in ion pairs.

4. Free Energy Profiles

QM/MM free energy calculations showed that the barrier for proton transfer from Asp61D1 to Glu310D2 is approximately 6 kcal mol⁻¹, with a reaction free energy change (ΔG) of about 3 kcal mol⁻¹. After YZ oxidation, the effective timescale for proton transfer is on the order of nanoseconds to microseconds.

Conclusion

This study elucidates the mechanism of stepwise deprotonation during light-driven water oxidation in the CL1 channel of PSII. Proton transfer is tightly controlled by redox-gated mechanisms that involve hydration transitions and ion pair conformational changes. The Glu65D1 gate modulates the proton transfer barrier in the CL1 channel, preventing proton backflow. Glu312D2 may serve as a local proton storage site. The findings suggest that several bioenergetic systems may employ electric field effects, ion pair conformational changes, and hydration transitions to control proton transfer reactions.

Research Highlights

1. Revealing the Proton Transfer Pathway: The study provides the first detailed description of the proton transfer pathway in the CL1 channel of PSII, highlighting the critical role of carboxylate chains in water-mediated proton transfer.
2. Discovery of Molecular Gating Mechanism: Glu65D1 is identified as a local molecular gate that controls proton transfer to the thylakoid lumen, preventing proton backflow.
3. Unveiling Electric Field Effects: The study, through simulations, reveals how the electric field triggered by YZ oxidation enhances proton transfer, offering new insights into the mechanism of proton transfer in bioenergetic systems.
4. Application of Multiscale Simulation Methods: The research combines classical MD simulations with QM/MM free energy calculations, providing a new methodological framework for studying proton transfer mechanisms in complex biological systems.

Research Significance

This study not only deepens the understanding of the water oxidation mechanism in PSII but also provides important theoretical insights into proton transfer mechanisms in other bioenergetic systems. The revealed electric field effects and molecular gating mechanisms may have broad applications in the study of bioenergetic systems, holding significant scientific and practical value.