Spectroscopic and Kinetic Studies of the Reaction of Bromopropanesulfonate with Methyl-coenzyme M Reductase

Ryan C. Kunz, Yih-Chern Horng, Stephen W. Ragsdale
2006 Journal of Biological Chemistry  
Methyl-coenzyme M reductase (MCR) catalyzes the final step of methanogenesis in which coenzyme B and methyl-coenzyme M are converted to methane and the heterodisulfide, CoMS-SCoB. MCR also appears to initiate anaerobic methane oxidation (reverse methanogenesis). At the active site of MCR is coenzyme F 430 , a nickel tetrapyrrole. This paper describes the reaction of the active MCR red1 state with the potent inhibitor, 3-bromopropanesulfonate (BPS; I 50 ‫؍‬ 50 nM) by UV-visible and EPR
more » ... e and EPR spectroscopy and by steady-state and rapid kinetics. BPS was shown to be an alternative substrate of MCR in an ionic reaction that is coenzyme B-independent and leads to debromination of BPS and formation of a distinct state ("MCR PS ") with an EPR signal that was assigned to a Ni(III)-propylsulfonate species (Hinderberger, D., Piskorski, R. P., Goenrich, M., Thauer, R. K., Schweiger, A., Harmer, J., and Jaun, B. (2006) Angew. Chem. Int. Ed. Engl. 45, 3602-3607). A similar EPR signal was generated by reacting MCR red1 with several halogenated sulfonate and carboxylate substrates. In rapid chemical quench experiments, the propylsulfonate ligand was identified by NMR spectroscopy and high performance liquid chromatography as propanesulfonic acid after protonolysis of the MCR PS complex. Propanesulfonate formation was also observed in steady-state reactions in the presence of Ti(III) citrate. Reaction of the alkylnickel intermediate with thiols regenerates the active MCR red1 state and eliminates the propylsulfonate group, presumably as the thioether. MCR PS is catalytically competent in both the generation of propanesulfonate and reformation of MCR red1 . These results provide evidence for the intermediacy of an alkylnickel species in the final step in anaerobic methane oxidation and in the initial step of methanogenesis. Downloaded from 8 The DFT calculations suggest that release of the methyl radical has a barrier of 19.5 kcal/mol, and formation of the bond between Ni(II) and the thiol sulfur of CoM has a barrier of 38.6 kcal/mol, yielding a net barrier for this step in the proposed reaction cycle of ϳ20 kcal/mol.
doi:10.1074/jbc.m606715200 pmid:16966321 fatcat:gr4ptoee4na6bakflvs56obeym