Resolving intermediates in biological proton-coupled electron transfer: A tyrosyl radical prior to proton movement

Abstract

The coupling of proton chemistry with redox reactions is important in many enzymes and is central to energy transduction in biology. However, the mechanistic details are poorly understood. Here, we have studied tyrosine oxidation, a reaction in which the removal of one electron from the amino acid is linked to the release of its phenolic proton. Using the unique photochemical properties of photosystem II, it was possible to oxidize the tyrosine at 1.8 K, a temperature at which proton and protein motions are limited. The state formed was detected by high magnetic field EPR as a high-energy radical intermediate trapped in an unprecedentedly electropositive environment. Warming of the protein allows this state to convert to a relaxed, stable form of the radical. The relaxation event occurs at 77 K and seems to involve proton migration and only a very limited movement of the protein. These reactions represent a stabilization process that prevents the backreaction and determines the reactivity of the radical.

Mimicking the electron transfer chain in photosystem II with a molecular triad thermodynamically capable of water oxidation

“…The combined transient absorption and fluorescence results therefore reveal that the formation (590 ps) of the PF 10 •þ -TCNP •− state is slower than its decay (12 ps). This order of rate constants is an example of inverted kinetics, where the rate constant for recombination is larger than that for charge separation, and transient signals can be inverted in amplitude relative to their usual appearance (24,25). A quantum yield of 77% is calculated for formation of PF 10 •þ -TCNP •− in 2.…”

“…The resulting oxidation of the Bi-PhOH unit dramatically enhances the acidity of the phenolic proton (Δ pKa ∼ 12) (25,26), which creates the chemical potential required for proton transfer to the hydrogen-bonded benzimidazole moiety (27)(28)(29)(30). Therefore, excitation of triad 1 is expected to lead to eventual formation of a charge-separated state characterized by cationic benzimidazole, a phenoxyl radical, a neutral PF 10 , and a reduced TCNP porphyrin (BiH þ -PhO • -PF 10 -TCNP •− ).…”

Mechanism of tyrosine D oxidation in Photosystem II

“…Remarkably, under these high-pH conditions, TyrD oxidation can occur at the temperature of liquid helium, indicating a barrier-less proton transfer (16,17).…”

“…The mechanistic model for TyrD oxidation proposed here may provide insights to help understand the remarkable speed-up in the rate of TyrD oxidation at pH values higher than 7.6 (11,16,17). In the absence of a crystal structure at this pH, the situation will inevitably be less clear-cut.…”

Site-Directed Mutagenesis ofThermosynechococcus elongatusPhotosystem II: The O2-Evolving Enzyme Lacking the Redox-Active Tyrosine D

“…Although the deduced amino acid sequences of D1 and D2 share a rather low homology (30%) (10,11), the important amino acid residues such as those that act as ligands to the cofactors are conserved. The central Chl molecules (P D1 and P D2 , Chl D1 and Chl D2 ), pheophytins (Pheo D1 and Pheo D2 ), plastoquinones (Q B and Q A ), peripheral Chls (Chl ZD1 and Chl ZD2 ), and two redox-active tyrosines (Tyr Z and Tyr D ) occupy symmetrical position on the D1 and D2 proteins and are hydrogen-bonded to D1-H190 and D2-H189, respectively (5)(6)(7)(12)(13)(14)(15), while the Mn 4 CaO 4 cluster is located in D1 (1)(2)(3)(4)(5)(6)(7).…”