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On the catalytic mechanism and activation of methyl coenzyme M reductase (MCR): Two sides of the same coin

Ryan C Kunz, University of Nebraska - Lincoln

Abstract

Methyl-coenzyme M reductase (MCR) from the methanogenic archaeon Methanothermobacter marburgensis catalyzes the final step in the biological synthesis of methane. Using coenzyme B (CoB7SH) as the two-electron donor, MCR reduces methyl-coenzyme M (methyl-SCoM2) to methane and the mixed disulfide, CoBS-S-CoM. MCR contains an essential redox-active nickel tetrahydrocorphin at its active site called coenzyme F 430; the active form of MCR (MCRred1) contains Ni(I)-F 430. The role of Ni(I)-F430 in the catalytic cycle is not well understood. Based on crystal structures and computational studies, two catalytic mechanisms have been put forth: an SN2-type mechanism with a methyl-Ni(III) intermediate (mechanism I), and a methyl radical based mechanism (mechanism II), respectively. In addition, the activation of this novel enzyme is of significant chemical and biological interest. Spectroscopic and computational studies have been used to study novel forms of the coenzyme, called F330 and F380, which are obtained by reducing F430 with sodium borohydride (NaBH4) or Ti(III) citrate, respectively. Furthermore, we describe the reaction of the MCRred1 state with the potent inhibitor, 3-bromopropanesulfonate (BPS) by kinetic and spectroscopic methods. BPS was shown to be an alternative substrate of MCR in an ionic reaction that is CoBSH independent and leads to debromination of BPS and formation of a distinct EPR-active state called “MCR PS” which is best described as a Ni(III)-propylsulfonate complex. A similar EPR signal was generated by reacting MCRred1 with several halogenated sulfonate and a series of brominated carboxylic acids, with chain lengths ranging from 4 to 16 carbons. Reaction of the alkylnickel intermediate with thiols regenerates the active MCRred1. Mass spectrometric evidence demonstrates thioether formation from this reaction. Interestingly, MCRPS can also be reductively activated with analogs of CoBSH such as, CoB8SH and CoB9SH. In this context, BPS is best described not as a competitive inhibitor or an irreversible inhibitor but as an alternative substrate. In a similar manner, treating MCRred1 with iodomethane generates a methyl-Ni(III) and, sets the foundation for testing the relevance of this species in catalysis. When reacted with methyl-SCoM and CoB6SH, the decay of MCRred1 is observed, and a new organic radical species is observed by EPR. ^

Subject Area

Biology, Molecular|Biology, Microbiology|Biophysics, General

Recommended Citation

Kunz, Ryan C, "On the catalytic mechanism and activation of methyl coenzyme M reductase (MCR): Two sides of the same coin" (2007). ETD collection for University of Nebraska - Lincoln. AAI3275069.
http://digitalcommons.unl.edu/dissertations/AAI3275069

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