Role of the [2Fe-2S]2+ cluster in biotin synthase: mutagenesis of the atypical metal ligand arginine 260.

Role of the [2Fe-2S]2+ cluster in biotin synthase: mutagenesis of the atypical metal ligand arginine 260. Research Abstract Details 

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Role of the [2Fe-2S]2+ cluster in biotin synthase: mutagenesis of the atypical metal ligand arginine 260. Abstract Text:

Robyn B Broach,Joseph T Jarrett,

Biotin synthase (BS) is an S-adenosylmethionine (AdoMet)-dependent radical enzyme that catalyzes the addition of sulfur to dethiobiotin. Like other AdoMet radical enzymes, BS contains a [4Fe-4S] cluster that is coordinated by a highly conserved CxxxCxxC sequence motif and by the methionyl amine and carboxylate of AdoMet. The close association of the [4Fe-4S]+ cluster with AdoMet facilitates reductive cleavage of the sulfonium and the generation of transient 5'-deoxyadenosyl radicals, which are then proposed to sequentially abstract hydrogen atoms from the substrate to produce carbon radicals at C9 and C6 of dethiobiotin. BS also contains a [2Fe-2S]2+ cluster located approximately 4-5 A from dethiobiotin, and we have proposed that a bridging sulfide of this cluster quenches the substrate radicals, leading to formation of the thiophane ring of biotin. In BS from Escherichia coli, the [2Fe-2S]2+ cluster is coordinated by cysteines 97, 128, and 188, and the atypical metal ligand, arginine 260. The evolutionary conservation of an arginine guanidinium as a metal ligand suggests a novel role for this residue in tuning the reactivity or stability of the [2Fe-2S]2+ cluster. In this work, we explore the effects of mutagenesis of Arg260 to Ala, Cys, His, and Met. Although perturbations in a number of characteristics of the [2Fe-2S]2+ cluster and the proteins are noted, the reconstituted enzymes have in vitro single-turnover activities that are 30-120% of that of the wild type. Further, in vivo expression of each mutant enzyme was sufficient to sustain growth of a bioB- mutant strain on dethiobiotin-supplemented medium, suggesting the enzymes were active and efficiently reconstituted by the in vivo iron-sulfur cluster (ISC) assembly system. Although we cannot exclude an as-yet-unidentified in vivo role in cluster repair or retention, we can conclude that Arg260 is not essential for the catalytic reaction of BS.

Role of the [2Fe-2S]2+ cluster in biotin synthase: mutagenesis of the atypical metal ligand arginine 260. Publishing Authors By Initials

RB Broach,JT Jarrett,

For similar enzymes and coenzymes: enzymes: transferases: sulfur group transferases: sulfurtransferases research abstracts see: enzymes and coenzymes: enzymes: transferases: sulfur group transferases: sulfurtransferases research

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Role of the [2Fe-2S]2+ cluster in biotin synthase: mutagenesis of the atypical metal ligand arginine 260. Journal Published:

PUBLICATION TYPE: Research Support, Non-U.S. Gov

Journal: Biochemistry

VOLUME: 45

Page Numbers: 14166-74

Journal Abbreviation: Biochemistry

ISSN: 0006-2960

DAY: 28

MONTH: Nov

YEAR: 2006

Role of the [2Fe-2S]2+ cluster in biotin synthase: mutagenesis of the atypical metal ligand arginine 260. Information

Number of References:

LANGUAGE: eng

NlmUniqueID: 370623

Role of the [2Fe-2S]2+ cluster in biotin synthase: mutagenesis of the atypical metal ligand arginine 260. Keywords Mesh Terms:

KEYWORDS: Sulfurtransferases

MESH TERMS: metabolism

Chemical & Substance for Abstract: Role of the [2Fe-2S]2+ cluster in biotin synthase: mutagenesis of the atypical metal ligand arginine 260. Information

Substance Name: biotin synthetase

Registry Number: EC 2.8.1.6

Grant and Affiliation Information for Role of the [2Fe-2S]2+ cluster in biotin synthase: mutagenesis of the atypical metal ligand arginine 260.

AFFILIATION: Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.

Country: United States

AGENCY: United States NIGMS

GRANT: R01 GM59175

ACRONYM: GM

MEDLINETA: Biochemistry

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