X-ray absorption spectroscopy of the corrinoid/iron-sulfur protein involved in acetyl coenzyme A synthesis by Clostridium thermoaceticum

Wirt, Michael D.; Kumar, Manoj; Ragsdale, Stephen W.; Chance, Mark R.

doi:10.1021/ja00059a007

Cited by 39 publications

(41 citation statements)

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“…The corrinoidiron-sulfur proteins are similar to the other corrinoid proteins in that their corrinoid is bound in the base-off constitution, but they appear to differ from these in that the lower axial coordination position in the cob(I1)amide oxidation state is not a histidine: the EPR spectrum of the corrinoid ironsulfur protein in the cob(I1)amide oxidation state exhibits a large cobalt hyperfine splitting, but it lacks superhyperfine interactions (Ragsdale et al, 1987). The absence of a lower axial coordination position was also shown by X-ray absorption spectroscopy (Wirt et al, 1993).…”

Section: Discussionmentioning

confidence: 62%

The Corrinoid‐Containing 23‐kDa Subunit MtrA of the Energy‐Conserving N⁵‐Methyltetrahydromethanopterin:coenzyme M Methyltransferase Complex from Methanobacterium Thermoautotrophicum

Harms

Thauer

1996

European Journal of Biochemistry

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Ni-Methy1tetrahydromethanopterin:coenzyme M methyltransferase (Mtr) from Methanobacterium thermoautotrophicum is a membrane-associated enzyme complex that catalyzes an energy-conserving, sodium ion translocating step in methanogenesis from H, and CO,. The complex is composed of eight different subunits, MtrA-H, one of which (MtrA) harbours a corrinoid as prosthetic group. In this study, we report the structural properties of MtrAl [des-(214-239)-MtrA], which is a deletion mutant of MtrA that lacks the last 25 C-terminal hydrophobic amino acids rendering the membrane protein soluble :(a) mtrAl was heterologously expressed in Escherichiu coli. Overexpression yielded a cytoplasmic protein which was purified approximately tenfold to apparent homogeneity. The purified protein was devoid of its corrinoid prosthetic group and not correctly folded as was evident from its electrophoretic mobility in SDS/PAGE. (b) Unfolding of MtrAl with guanidine/HCl and refolding in the presence of cobalamin resulted in the formation of the correctly folded MtrAl holoprotein that contained tightly bound cob(I1)-alamin; the rate of reconstitution was highest when the refolding proceeded in the presence of titanium(II1) citrate, which suggested that cob(1)alamin is the corrinoid species that binds to the apoprotein. (c) EPR spectra of the cob(I1)alamin-containing holoprotein differentially labelled with 14N (nuclear spin 1) and "N (nuclear spin 1/2) revealed that the corrinoid is bound to MtrAl in the base-off form and that the Co(I1) of the prosthetic group is coordinated by a histidine residue of the apoprotein. The results are interpreted with respect to the mechanism of energy conservation by the MtrA-H complex.

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Section: Discussionmentioning

confidence: 62%

The Corrinoid‐Containing 23‐kDa Subunit MtrA of the Energy‐Conserving N⁵‐Methyltetrahydromethanopterin:coenzyme M Methyltransferase Complex from Methanobacterium Thermoautotrophicum

Harms

Thauer

1996

European Journal of Biochemistry

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“…In the M. thermoaceticum MTHF:CFeSP methyltransferase involved in the Wood-Ljungdahl pathway, the inactive Co(II) state of the CFeSP is already "base-off " in the resting enzyme, which represents a "ready" state for electron transfer to form a four-coordinate Co(I) state (Harder et al, 1989;Ragsdale et al, 1987;Stich et al, 2006;Wirt et al, 1993Wirt et al, , 1995. The direct electron donor is the [4Fe-4S] cluster of the AcsC subunit of the CFeSP (Menon and Ragsdale, 1999).…”

Section: B Generation and Maintenance Of The Active Co(i) State Of B 12mentioning

confidence: 99%

Catalysis of Methyl Group Transfers Involving Tetrahydrofolate and B12

Ragsdale

2008

Vitamins &Amp; Hormones

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This review focuses on the reaction mechanism of enzymes that use B(12) and tetrahydrofolate (THF) to catalyze methyl group transfers. It also covers the related reactions that use B(12) and tetrahydromethanopterin (THMPT), which is a THF analog used by archaea. In the past decade, our understanding of the mechanisms of these enzymes has increased greatly because the crystal structures for three classes of B(12)-dependent methyltransferases have become available and because biophysical and kinetic studies have elucidated the intermediates involved in catalysis. These steps include binding of the cofactors and substrates, activation of the methyl donors and acceptors, the methyl transfer reaction itself, and product dissociation. Activation of the methyl donor in one class of methyltransferases is achieved by an unexpected proton transfer mechanism. The cobalt (Co) ion within the B(12) macrocycle must be in the Co(I) oxidation state to serve as a nucleophile in the methyl transfer reaction. Recent studies have uncovered important principles that control how this highly reducing active state of B(12) is generated and maintained.

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“…16,17 Yet, on the basis of our recent computational studies on a series of Co 2+ Cbi + models with varying Co-OH 2 bond lengths, lower axial ligand dissociation should drastically alter the energies of the Co 3d-based ligand field (LF) transitions and the EPR parameters (e.g., g-values and 59 Co hyperfine coupling constants). 27 However, no such dramatic alterations are actually observed in the case of asisolated Co 2+ CFeSP, thus arguing against the presence of a four-coordinate Co 2+ corrinoid species in this enzyme.…”

Section: Reactivation Of Co 2+ Cfespmentioning

confidence: 99%

“…Nonetheless, a subsequent detailed analysis of the pre-edge features in the X-ray absorption (XAS) spectrum of the Co 2+ CFeSP continued to suggest that the Co 2+ corrinoid cofactor exists in an unprecedented four-coordinate square-planar geometry. 16,18 Numerous thermodynamic studies of alkylated corrinoid model complexes have shown that the nature of the lower ligand has a considerable influence on both the mode (i.e., homolytic vs heterolytic) and the rate of Co-C bond cleavage. [19][20][21][22][23] Consequently, the lower axial ligand is expected to play a key role in controlling both the reaction of Co 1+ CFeSP with CH 3 -H 4 folate and the transfer of the methyl cation from Me-Co 3+ CFeSP to the ACS A-cluster, and it is therefore of considerable interest to unambiguously establish the binding scheme of the corrinoid cofactor in CFeSP.…”

Section: Introductionmentioning

confidence: 99%

Spectroscopic Studies of the Corrinoid/Iron−Sulfur Protein from Moorella thermoacetica

et al. 2006

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Methyl transfer reactions are important in a number of biochemical pathways. An important class of methyltransferases uses the cobalt cofactor cobalamin, which receives a methyl group from an appropriate methyl donor protein to form an intermediate organometallic methyl-Co bond that subsequently is cleaved by a methyl acceptor. Control of the axial ligation state of cobalamin influences both the mode (i.e., homolytic vs heterolytic) and the rate of Co-C bond cleavage. Here we have studied the axial ligation of a corrinoid iron-sulfur protein (CFeSP) that plays a key role in energy generation and cell carbon synthesis by anaerobic microbes, such as methanogenic archaea and acetogenic bacteria. This protein accepts a methyl group from methyltetrahydrofolate forming Me-Co 3+ CFeSP that then donates a methyl cation (Me) from Me-Co 3+ CFeSP to a nickel site on acetyl-CoA synthase. To unambiguously establish the binding scheme of the corrinoid cofactor in the CFeSP, we have combined resonance Raman, magnetic circular dichroism, and EPR spectroscopic methods with computational chemistry. Our results clearly demonstrate that the MeCo 3+ and Co 2+ states of the CFeSP have an axial water ligand like the free MeCbi + and Co 2+ Cbi + cofactors; however, the Co-OH 2 bond length is lengthened by about 0.2 Å for the protein-bound cofactor. Elongation of the Co-OH 2 bond of the CFeSP-bound cofactor is proposed to make the cobalt center more "Co 1+ -like", a requirement to facilitate heterolytic Co-C bond cleavage.

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X-ray absorption spectroscopy of the corrinoid/iron-sulfur protein involved in acetyl coenzyme A synthesis by Clostridium thermoaceticum

Cited by 39 publications

References 1 publication

The Corrinoid‐Containing 23‐kDa Subunit MtrA of the Energy‐Conserving N⁵‐Methyltetrahydromethanopterin:coenzyme M Methyltransferase Complex from Methanobacterium Thermoautotrophicum

The Corrinoid‐Containing 23‐kDa Subunit MtrA of the Energy‐Conserving N⁵‐Methyltetrahydromethanopterin:coenzyme M Methyltransferase Complex from Methanobacterium Thermoautotrophicum

Catalysis of Methyl Group Transfers Involving Tetrahydrofolate and B12

Spectroscopic Studies of the Corrinoid/Iron−Sulfur Protein from Moorella thermoacetica

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X-ray absorption spectroscopy of the corrinoid/iron-sulfur protein involved in acetyl coenzyme A synthesis by Clostridium thermoaceticum

Cited by 39 publications

References 1 publication

The Corrinoid‐Containing 23‐kDa Subunit MtrA of the Energy‐Conserving N5‐Methyltetrahydromethanopterin:coenzyme M Methyltransferase Complex from Methanobacterium Thermoautotrophicum

The Corrinoid‐Containing 23‐kDa Subunit MtrA of the Energy‐Conserving N5‐Methyltetrahydromethanopterin:coenzyme M Methyltransferase Complex from Methanobacterium Thermoautotrophicum

Catalysis of Methyl Group Transfers Involving Tetrahydrofolate and B12

Spectroscopic Studies of the Corrinoid/Iron−Sulfur Protein from Moorella thermoacetica

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The Corrinoid‐Containing 23‐kDa Subunit MtrA of the Energy‐Conserving N⁵‐Methyltetrahydromethanopterin:coenzyme M Methyltransferase Complex from Methanobacterium Thermoautotrophicum

The Corrinoid‐Containing 23‐kDa Subunit MtrA of the Energy‐Conserving N⁵‐Methyltetrahydromethanopterin:coenzyme M Methyltransferase Complex from Methanobacterium Thermoautotrophicum