How Do Metalloproteins Tame the Fenton Reaction and Utilize •OH Radicals in Constructive Manners?

CONSPECTUS: This Account describes the manner whereby nature controls the Fenton-type reaction of O-O homolysis of hydrogen peroxide and harnesses it to carry out various useful oxidative transformations in metalloenzymes. H2O2 acts as the cosubstrate for the heme-dependent peroxidases, P450BM3, P450(SP alpha), P450(BS beta), and the P450 decarboxylase OleT, as well as the nonheme enzymes HppE and the copper-dependent lytic polysaccharide monooxygenases (LPMOs). Whereas heme perox-idases use the Poulos-Kraut heterolytic mechanism for H2O2 activation, some heme enzymes prefer the alternative Fenton-type mechanism, which produces center dot OH radical intermediates. The fate of the center dot OH radical is controlled by the protein environment, using tight H-bonding networks around H2O2. The so-generated center dot OH radical is constrained by the surrounding H-bonding interactions, the orientation of which is targeted to perform H-abstraction from the Fe(III)-OH group and thereby leading to the formation of the active species, called Compound I (Cpd I), Por(+center dot)Fe(IV)= O, which performs oxidation of the substrate. Alternatively, for the nonheme HppE enzyme, the O-O homolysis catalyzed by the resting state Fe(II) generates an Fe(III)-OH species that effectively constrains the center dot OH radical species by a tight H-bonding network. The so-formed H-bonded center dot OH radical acts directly as the oxidant, since it is oriented to perform H-abstraction from the C-H bond of the substrate (S)-2-HPP. The Fenton-type H2O2 activation is strongly suggested by computations to occur also in copper-dependent LPMOs and pMMO. In LPMOs, the Cu(I)-catalyzed O-O homolysis of the H2O2 cosubstrate generates an center dot OH radical that abstracts a hydrogen atom from Cu(II)-OH and forms thereby the active species of the enzyme, Cu(II)-O center dot. Such Fenton-type O-O activation can be shared by both the O-2-dependent activations of LPMOs and pMMOs, in which the O-2 cosubstrate may be reduced to H2O2 by external reductants. Our studies show that, generally, the H2O2 activation is highly dependent on the protein environment, as well as on the presence/absence of substrates. Since H2O2 is a highly flexible and hydrophilic molecule, the absence of suitable substrates may lead to unproductive binding or even to the release of H2O2 from the active site, as has been suggested in P450cam and LPMOs, whereas the presence of the substrate seems to play a role in steering a Fenton-type H2O2 activation. In the absence of a substrate, the hydrophilic active site of P450BM3 disfavors the binding and activation of H2O2 and protects thereby the enzyme from the damage by the Fenton reaction. Due to the distinct coordination and reaction environment, the Fenton-type H2O2 activation mechanism by enzymes differs from the reaction in synthetic systems. In nonenzymatic reactions, the H-bonding networks are quite dynamic and flexible and the reactivity of H2O2 is not strategically constrained as in the enzymatic environment. As such, our Account describes the controlled Fenton-type mechanism in metalloenzymes, and the role of the protein environment in constraining the center dot OH radical against oxidative damage, while directing it to perform useful oxidative transformations.

Automated summary

This Account describes how nature controls the Fenton-type reaction of hydrogen peroxide and utilizes it for various oxidative transformations in metalloenzymes. The protein environment plays a crucial role in constraining and directing the ·OH radical, which is generated from O-O homolysis of hydrogen peroxide, to perform useful oxidative transformations. The presence or absence of substrates and the specific protein environment greatly influence the activation of hydrogen peroxide.