The L-form of N-7-mercaptoheptanoyl-O-phosphothreonine is the enantiomer active as component B in methyl-CoM reduction to methane

AbstractThe reduction of methyl-CoM to methane in methanogenic bacteria is dependent on a low-Mr, heat-stable compound designated component B, the structure of which has recently been assigned as N-7-mercaptoheptanoyl-O-phosphothreonine. We report here that only the enantiomer derived from O-phospho-L-threonine has cofactor activity. N-7-Mercaptoheptanoyl-O-phospho-D-threonine was neither active nor inhibitory

7-Mercaptoheptanoylthreonine phosphate functions as component B in ATP-independent methane formation from methyl-CoM with reduced cobalamin as electron donor

AbstractPurified methyl-CoM reductase of Methanobacterium thermoautotrophicum (strain Marburg) catalyzed the reduction of methyl-CoM to methane with reduced cobalamin, when either synthetic 7-mercaptoheptanoyl-threonine phosphate (HS-HTP) or naturally occurring component B was present. With both compounds the same maximal specific acitivity was obtained and ATP was neither required nor stimulatory. These findings indicate that HS-HTP functions as component B and do not support the idea that HS-HT is only active in an adenosine monophosphorylated form

Cloning and characterization of the methyl coenzyme M reductase genes from Methanobacterium thermoautotrophicum.

The genes coding for methyl coenzyme M reductase were cloned from a genomic library of Methanobacterium thermoautotrophicum Marburg into Escherichia coli by using plasmid expression vectors. When introduced into E. coli, the reductase genes were expressed, yielding polypeptides identical in size to the three known subunits of the isolated enzyme, alpha, beta, and gamma. The polypeptides also reacted with the antibodies raised against the respective enzyme subunits. In M. thermoautotrophicum, the subunits are encoded by a gene cluster whose transcript boundaries were mapped. Sequence analysis revealed two more open reading frames of unknown function located between two of the methyl coenzyme M reductase genes

The magnetic properties of the nickel cofactor F430 in the enzyme methyl-coenzyme M reductase of Methanobacterium thermoautotrophicum.

Cofactor 430 of methyl-coenzyme M reductase from Methanobacterium thermoautotrophicum was studied in both the extracted form in aqueous solution and protein-bound by using low-temperature magnetic-circular-dichroism spectroscopy. In both forms the nickel was present as high-spin paramagnetic nickel(II), spin S = 1, subject to almost equal zero-field splitting (cofactor F430, D = +9.0 cm-1, E/D = 0; methyl-coenzyme M reductase, D = +8.5 cm-1, [E/D[ = 0.2). This suggests identical axial co-ordination by oxygen ligand(s) both in aqueous cofactor F430 and in the investigated state of the protein