Dissecting Electronic-Structural Transitions in the Nitrogenase MoFe Protein P-Cluster during Reduction

Area of Science:

• Biochemistry
• Bioinorganic Chemistry
• Enzyme Catalysis

Background:

• The [8Fe-7S] P-cluster in nitrogenase MoFe protein is vital for electron transfer from the Fe protein during ammonia synthesis.
• The P-cluster cycles through oxidation states (P^N, P⁺, P²⁺), with the P^N↔P⁺ transition being critical for nitrogenase complex activity.

Purpose of the Study:

• To investigate the intermediate steps in the formation of the P⁺ oxidation state during electron transfer.
• To understand the coupling between spin-state transitions and oxidation state changes in the P-cluster.

Main Methods:

• Photochemical reduction of MoFe protein at low temperatures (231-263 K) to trap intermediates.
• Electron Paramagnetic Resonance (EPR) spectroscopy to analyze trapped intermediates.
• Kinetic analysis of EPR signal changes during dark annealing.

Main Results:

• Illumination of MoFe protein with CdS nanocrystals induced P-cluster reduction (P²⁺).
• Three distinct EPR signals (S=1/2 axial, S=1/2 rhombic, S=7/2 high-spin) were observed.
• Dark annealing showed a decrease in axial and high-spin signals, with a concurrent increase in the rhombic signal, indicating their intermediate roles.

Conclusions:

• The axial and high-spin EPR signals represent transient intermediates in the formation of the P⁺ resting state.
• Spin-state changes are intrinsically linked to oxidation state fluctuations within the P-cluster during electron transfer.