Characterization of alkyl-nickel adducts generated by reaction of methyl-coenzyme m reductase with brominated acids.
Methyl-coenzyme M reductase (MCR) from methanogenic archaea catalyzes the final step in the biological synthesis of methane. Using coenzyme B (CoBSH) as the two-electron donor, MCR reduces methyl-coenzyme M (methyl-SCoM) to methane and the mixed disulfide, CoB-S-S-CoM. MCR contains coenzyme F430, an essential redox-active nickel tetrahydrocorphin, at its active site. The active form of MCR (MCRred1) contains Ni(I)-F430. When 3-bromopropane sulfonate (BPS) is incubated with MCRred1, an alkyl-Ni(III) species is formed that elicits the MCRPS EPR signal. Here we used EPR and UV-visible spectroscopy and transient kinetics to study the reaction between MCR from Methanothermobacter marburgensis and a series of brominated carboxylic acids, with carbon chain lengths of 4-16. All of these compounds give rise to an alkyl-Ni intermediate with an EPR signal similar to that of the MCRPS species. Reaction of the alkyl-Ni(III) adduct, formed from brominated acids with eight or fewer total carbons, with HSCoM as nucleophile at pH 10.0 results in the formation of a thioether coupled to regeneration of the active MCRred1 state. When reacted with 4-bromobutyrate, MCRred1 forms the alkyl-Ni(III) MCRXA state and then, surprisingly, undergoes "self-reactivation" to regenerate the Ni(I) MCRred1 state and a bromocarboxy ester. The results demonstrate an unexpected reactivity and flexibility of the MCR active site in accommodating a broad range of substrates, which act as molecular rulers for the substrate channel in MCR.
Dey
Mishtu
M
Department of Biochemistry, University of Nebraska, Lincoln, Nebraska 68588, USA.
Kunz
Ryan C
RC
Lyons
Derek M
DM
Ragsdale
Stephen W
SW
D017738:Alkanesulfonic Acids / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D002264:Carboxylic Acids / Q000737:chemistry / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy; D007700:Kinetics; D017013:Methanobacteriaceae / Q000201:enzymology; D008956:Models, Chemical; D015394:Molecular Structure; D009532:Nickel / Q000737:chemistry* / Q000378:metabolism; D010088:Oxidoreductases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D011485:Protein Binding; D013056:Spectrophotometry, Ultraviolet
C495861:3-bromopropanesulfonate; D017738:Alkanesulfonic Acids; D002264:Carboxylic Acids; D009532:Nickel; D010088:Oxidoreductases; C021572:methyl coenzyme M reductase