Showing papers by "Reiner Hedderich published in 2001"
TL;DR: In EPR spectroscopic studies with the enzyme from Methanothermobacter marburgensis, a unique paramagnetic species is identified that is formed upon reaction of the oxidized enzyme with H-S-CoM in the absence of H- S-CoB.
Abstract: Heterodisulfide reductase (Hdr) from methanogenic archaea is an iron–sulfur protein that catalyses the reversible reduction of the heterodisulfide (CoM-S-S-CoB) of the methanogenic thiol coenzymes, coenzyme M (H-S-CoM) and coenzyme B (H-S-CoB). In EPR spectroscopic studies with the enzyme from Methanothermobacter marburgensis, we have identified a unique paramagnetic species that is formed upon reaction of the oxidized enzyme with H-S-CoM in the absence of H-S-CoB. This paramagnetic species can be reduced in a one-electron step with a midpoint-potential of −185 mV but not further oxidized. A broadening of the EPR signal in the 57Fe-enriched enzyme indicates that it is at least partially iron based. The g values (gxyz= 2.013, 1.991 and 1.938) and the midpoint potential argue against a conventional [2Fe−2S]+, [3Fe−4S]+, [4Fe−4S]+ or [4Fe−4S]3+ cluster. This species reacts with H-S-CoB to form an EPR silent form. Hence, we propose that only a half reaction is catalysed in the presence of H-S-CoM and that a reaction intermediate is trapped. This reaction intermediate is thought to be a [4Fe−4S]3+ cluster that is coordinated by one of the cysteines of a nearby active-site disulfide or by the sulfur of H-S-CoM. A paramagnetic species with similar EPR properties was also identified in Hdr from Methanosarcina barkeri.
48 citations
TL;DR: The rapid and selective isolation by chloroform/methanol offers a new way to obtain the large quantities of material required for biochemical analyses and molecular and biochemical data suggest that the proteolipid from M. thermoautotrophicus is a duplication of the 8-kDa proteolIPid usually present in other archaea, but it retained the conserved glutamate involved in proton translocation in every copy.
Abstract: Chloroform/methanol was applied to cytoplasmic membranes of the thermophilic methanogens Methanothermobacter thermoautotrophicus and Methanothermobacter marburgensis as well as to the mesophile Methanosarcina mazei Go1. In any case, the chloroform/methanol extraction yielded only two proteins, subunit D (MtrD) of the Na+-translocating methyltetrahydromethanopterin:coenzyme M methyltransferase and the proteolipid of the A1A0 ATPase. Both polypeptides are assumed to be directly involved in ion translocation in their respective enzymes, but have not been studied in detail due to lack of simple isolation procedures. The rapid and selective isolation by chloroform/methanol offers a new way to obtain the large quantities of material required for biochemical analyses. As a first result, molecular and biochemical data suggest that the proteolipid from M. thermoautotrophicus is a duplication of the 8-kDa proteolipid usually present in other archaea, but it retained the conserved glutamate involved in proton translocation in every copy. This is the first 16-kDa proteolipid found in archaea.
17 citations