Coordination of adenosylmethionine to a unique iron site of the [4Fe-4S] of pyruvate formate-lyase activating enzyme: A Mössbauer spectroscopic study

Carsten Krebs, William E. Broderick, Timothy F. Henshaw, Joan B. Broderick, Boi Hanh Huynh

Research output: Contribution to journalArticle

106 Citations (Scopus)

Abstract

Pyruvate formate-lyase activating enzyme (PFL-AE) generates the catalytically essential glycyl radical of PFL. It is a member of the so-called "radical-SAM superfamily" of enzymes that use a [4Fe-4S] cluster and S-adenosylmethionine (AdoMet or SAM) to catalyze diverse radical-mediated reactions. Evidence suggests that this class of enzymes operate by common initial steps involving the generation of an AdoMet-derived adenosyl radical intermediate, of which the mechanism remains unresolved. The three-cysteine CX3CX2C cluster-binding motif common to all members of this superfamily suggests a unique Fe site in the [4Fe-4S] cluster, which presumably interacts with AdoMet to effect the reductive cleavage and radical generation. Here we employ a dual-iron-isotope (56Fe/57Fe) approach to demonstrate the existence of a unique Fe site in the [4Fe-4S] cluster of PFL-AE by Mössbauer spectroscopy. Coordination of AdoMet to this unique Fe site was made evident by the observation of a substantial increase in the isomer shift (δ) of the Mössbauer spectrum associated with the unique Fe site: δ = 0.42 mm/s in the absence of AdoMet increases to δ = 0.72 mm/s in the presence of AdoMet. Further, the Mössbauer data show that the binding of AdoMet to the unique Fe site occurs in the [4Fe-4S]2+ state, prior to the injection of the reducing equivalent required for catalysis. This observation indicates that AdoMet coordination is a necessary prerequisite to adenosyl radical generation.

Original languageEnglish (US)
Pages (from-to)912-913
Number of pages2
JournalJournal of the American Chemical Society
Volume124
Issue number6
DOIs
StatePublished - Feb 12 2002

Fingerprint

S-Adenosylmethionine
Iron
Enzymes
Isomers
Catalysis
Isotopes
Iron Isotopes
Spectroscopy
pyruvate formate-lyase activating enzyme
Cysteine
Spectrum Analysis
Observation
Injections

All Science Journal Classification (ASJC) codes

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry

Cite this

@article{dee08d7af66e4ba38f0549a3de06cb34,
title = "Coordination of adenosylmethionine to a unique iron site of the [4Fe-4S] of pyruvate formate-lyase activating enzyme: A M{\"o}ssbauer spectroscopic study",
abstract = "Pyruvate formate-lyase activating enzyme (PFL-AE) generates the catalytically essential glycyl radical of PFL. It is a member of the so-called {"}radical-SAM superfamily{"} of enzymes that use a [4Fe-4S] cluster and S-adenosylmethionine (AdoMet or SAM) to catalyze diverse radical-mediated reactions. Evidence suggests that this class of enzymes operate by common initial steps involving the generation of an AdoMet-derived adenosyl radical intermediate, of which the mechanism remains unresolved. The three-cysteine CX3CX2C cluster-binding motif common to all members of this superfamily suggests a unique Fe site in the [4Fe-4S] cluster, which presumably interacts with AdoMet to effect the reductive cleavage and radical generation. Here we employ a dual-iron-isotope (56Fe/57Fe) approach to demonstrate the existence of a unique Fe site in the [4Fe-4S] cluster of PFL-AE by M{\"o}ssbauer spectroscopy. Coordination of AdoMet to this unique Fe site was made evident by the observation of a substantial increase in the isomer shift (δ) of the M{\"o}ssbauer spectrum associated with the unique Fe site: δ = 0.42 mm/s in the absence of AdoMet increases to δ = 0.72 mm/s in the presence of AdoMet. Further, the M{\"o}ssbauer data show that the binding of AdoMet to the unique Fe site occurs in the [4Fe-4S]2+ state, prior to the injection of the reducing equivalent required for catalysis. This observation indicates that AdoMet coordination is a necessary prerequisite to adenosyl radical generation.",
author = "Carsten Krebs and Broderick, {William E.} and Henshaw, {Timothy F.} and Broderick, {Joan B.} and Huynh, {Boi Hanh}",
year = "2002",
month = "2",
day = "12",
doi = "10.1021/ja017562i",
language = "English (US)",
volume = "124",
pages = "912--913",
journal = "Journal of the American Chemical Society",
issn = "0002-7863",
publisher = "American Chemical Society",
number = "6",

}

Coordination of adenosylmethionine to a unique iron site of the [4Fe-4S] of pyruvate formate-lyase activating enzyme : A Mössbauer spectroscopic study. / Krebs, Carsten; Broderick, William E.; Henshaw, Timothy F.; Broderick, Joan B.; Huynh, Boi Hanh.

In: Journal of the American Chemical Society, Vol. 124, No. 6, 12.02.2002, p. 912-913.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Coordination of adenosylmethionine to a unique iron site of the [4Fe-4S] of pyruvate formate-lyase activating enzyme

T2 - A Mössbauer spectroscopic study

AU - Krebs, Carsten

AU - Broderick, William E.

AU - Henshaw, Timothy F.

AU - Broderick, Joan B.

AU - Huynh, Boi Hanh

PY - 2002/2/12

Y1 - 2002/2/12

N2 - Pyruvate formate-lyase activating enzyme (PFL-AE) generates the catalytically essential glycyl radical of PFL. It is a member of the so-called "radical-SAM superfamily" of enzymes that use a [4Fe-4S] cluster and S-adenosylmethionine (AdoMet or SAM) to catalyze diverse radical-mediated reactions. Evidence suggests that this class of enzymes operate by common initial steps involving the generation of an AdoMet-derived adenosyl radical intermediate, of which the mechanism remains unresolved. The three-cysteine CX3CX2C cluster-binding motif common to all members of this superfamily suggests a unique Fe site in the [4Fe-4S] cluster, which presumably interacts with AdoMet to effect the reductive cleavage and radical generation. Here we employ a dual-iron-isotope (56Fe/57Fe) approach to demonstrate the existence of a unique Fe site in the [4Fe-4S] cluster of PFL-AE by Mössbauer spectroscopy. Coordination of AdoMet to this unique Fe site was made evident by the observation of a substantial increase in the isomer shift (δ) of the Mössbauer spectrum associated with the unique Fe site: δ = 0.42 mm/s in the absence of AdoMet increases to δ = 0.72 mm/s in the presence of AdoMet. Further, the Mössbauer data show that the binding of AdoMet to the unique Fe site occurs in the [4Fe-4S]2+ state, prior to the injection of the reducing equivalent required for catalysis. This observation indicates that AdoMet coordination is a necessary prerequisite to adenosyl radical generation.

AB - Pyruvate formate-lyase activating enzyme (PFL-AE) generates the catalytically essential glycyl radical of PFL. It is a member of the so-called "radical-SAM superfamily" of enzymes that use a [4Fe-4S] cluster and S-adenosylmethionine (AdoMet or SAM) to catalyze diverse radical-mediated reactions. Evidence suggests that this class of enzymes operate by common initial steps involving the generation of an AdoMet-derived adenosyl radical intermediate, of which the mechanism remains unresolved. The three-cysteine CX3CX2C cluster-binding motif common to all members of this superfamily suggests a unique Fe site in the [4Fe-4S] cluster, which presumably interacts with AdoMet to effect the reductive cleavage and radical generation. Here we employ a dual-iron-isotope (56Fe/57Fe) approach to demonstrate the existence of a unique Fe site in the [4Fe-4S] cluster of PFL-AE by Mössbauer spectroscopy. Coordination of AdoMet to this unique Fe site was made evident by the observation of a substantial increase in the isomer shift (δ) of the Mössbauer spectrum associated with the unique Fe site: δ = 0.42 mm/s in the absence of AdoMet increases to δ = 0.72 mm/s in the presence of AdoMet. Further, the Mössbauer data show that the binding of AdoMet to the unique Fe site occurs in the [4Fe-4S]2+ state, prior to the injection of the reducing equivalent required for catalysis. This observation indicates that AdoMet coordination is a necessary prerequisite to adenosyl radical generation.

UR - http://www.scopus.com/inward/record.url?scp=0037065661&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0037065661&partnerID=8YFLogxK

U2 - 10.1021/ja017562i

DO - 10.1021/ja017562i

M3 - Article

C2 - 11829592

AN - SCOPUS:0037065661

VL - 124

SP - 912

EP - 913

JO - Journal of the American Chemical Society

JF - Journal of the American Chemical Society

SN - 0002-7863

IS - 6

ER -