Methyl‐coenzyme‐M reductase from Methanobacterium thermoautotrophicum (strain Marburg)

Abstract

Methyl‐coenzyme‐M reductase from Methanobacterium thermoautotrophicum (strain Marburg) was purified to a stage where, besides the α, β and γ subunits, no additional polypeptides were detectable in the preparation. Under appropriate conditions the enzyme was found to catalyze the reduction of methyl‐CoM with 7‐mercaptoheptanoylthreonine phosphate (H‐S‐HTP) to CH₄ at a specific rate of 2.5 μmol · min⁻¹· mg protein⁻¹. This finding contradicts a recent report that methyl‐CoM reductase is only active when some contaminating proteins are present. The two polypeptides encoded by the open reading frames ORF₁ and ORF₂ of the methyl‐CoM reductase transcription unit did not co‐purify with the α, β and γ subunits. They were neither required nor did they stimulate the activity under the assay conditions. 3‐Bromopropanesulfonate (apparent K_i= 0.05 μM) and 2‐azidoethanesulfonate (apparent K_i= 1 μM) were found to be two new competitive inhibitors of methyl‐CoM reductase. Both inhibitors were considerably more effective than the “classical” 2‐bromoethanesulfonate (apparent K_i= 4 μM).