A new mode of B12 binding and the direct participation of a potassium ion in enzyme catalysis: X-ray structure of diol dehydratase

Naoki Shibata, Jun Masuda, Takamasa Tobimatsu, Tetsuo Toraya, Kyoko Suto, Yukio Morimoto, Noritake Yasuoka

Research output: Contribution to journalArticle

199 Citations (Scopus)

Abstract

Background: Diol dehydratase is an enzyme that catalyzes the adenosylcobalamin (coenzyme B12) dependent conversion of 1,2-diols to the corresponding aldehydes. The reaction initiated by homolytic cleavage of the cobalt-carbon bond of the coenzyme proceeds by a radical mechanism. The enzyme is an (α2β2γ2 heterooligomer and has an absolute requirement for a potassium ion for catalytic activity. The crystal structure analysis of a diol dehydratase-cyanocobalamin complex was carried out in order to help understand the mechanism of action of this enzyme. Results: The three- dimensional structure of diol dehydratase in complex with cyanocobalamin was determined at 2.2 Å, resolution. The enzyme exists as a dimer of heterotrimers (αβγ)2. The cobalamin molecule is bound between the α and β subunits in the 'base-on' mode, that is, 5,6-dimethylbenzimidazole of the nucleotide moiety coordinates to the cobalt atom in the lower axial position. The α subunit includes a (β/α)8 barrel. The substrate, 1,2-propanediol, and an essential potassium ion are deeply buried inside the barrel. The two hydroxyl groups of the substrate coordinate directly to the potassium ion. Conclusions: This is the first crystallographic indication of the 'base-on' mode of cobalamin binding. An unusually long cobalt-base bond seems to favor homolytic cleavage of the cobalt-carbon bond and therefore to favor radical enzyme catalysis. Reactive radical intermediates can be protected from side reactions by spatial isolation inside the barrel. On the basis of unique direct interactions between the potassium ion and the two hydroxyl groups of the substrate, direct participation of a potassium ion in enzyme catalysis is strongly suggested.

Original languageEnglish
Pages (from-to)997-1008
Number of pages12
JournalStructure
Volume7
Issue number8
DOIs
Publication statusPublished - Aug 15 1999

Fingerprint

Propanediol Dehydratase
Catalysis
Potassium
X-Rays
Ions
Vitamin B 12
Cobalt
Enzymes
Carbon
Propylene Glycol
Coenzymes
Aldehydes
Hydroxyl Radical
Nucleotides

Keywords

  • B enzyme
  • Diol dehydratase
  • Radicals
  • Reaction mechanism
  • TIM barrel

ASJC Scopus subject areas

  • Molecular Biology
  • Structural Biology

Cite this

A new mode of B12 binding and the direct participation of a potassium ion in enzyme catalysis : X-ray structure of diol dehydratase. / Shibata, Naoki; Masuda, Jun; Tobimatsu, Takamasa; Toraya, Tetsuo; Suto, Kyoko; Morimoto, Yukio; Yasuoka, Noritake.

In: Structure, Vol. 7, No. 8, 15.08.1999, p. 997-1008.

Research output: Contribution to journalArticle

Shibata, Naoki ; Masuda, Jun ; Tobimatsu, Takamasa ; Toraya, Tetsuo ; Suto, Kyoko ; Morimoto, Yukio ; Yasuoka, Noritake. / A new mode of B12 binding and the direct participation of a potassium ion in enzyme catalysis : X-ray structure of diol dehydratase. In: Structure. 1999 ; Vol. 7, No. 8. pp. 997-1008.
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abstract = "Background: Diol dehydratase is an enzyme that catalyzes the adenosylcobalamin (coenzyme B12) dependent conversion of 1,2-diols to the corresponding aldehydes. The reaction initiated by homolytic cleavage of the cobalt-carbon bond of the coenzyme proceeds by a radical mechanism. The enzyme is an (α2β2γ2 heterooligomer and has an absolute requirement for a potassium ion for catalytic activity. The crystal structure analysis of a diol dehydratase-cyanocobalamin complex was carried out in order to help understand the mechanism of action of this enzyme. Results: The three- dimensional structure of diol dehydratase in complex with cyanocobalamin was determined at 2.2 {\AA}, resolution. The enzyme exists as a dimer of heterotrimers (αβγ)2. The cobalamin molecule is bound between the α and β subunits in the 'base-on' mode, that is, 5,6-dimethylbenzimidazole of the nucleotide moiety coordinates to the cobalt atom in the lower axial position. The α subunit includes a (β/α)8 barrel. The substrate, 1,2-propanediol, and an essential potassium ion are deeply buried inside the barrel. The two hydroxyl groups of the substrate coordinate directly to the potassium ion. Conclusions: This is the first crystallographic indication of the 'base-on' mode of cobalamin binding. An unusually long cobalt-base bond seems to favor homolytic cleavage of the cobalt-carbon bond and therefore to favor radical enzyme catalysis. Reactive radical intermediates can be protected from side reactions by spatial isolation inside the barrel. On the basis of unique direct interactions between the potassium ion and the two hydroxyl groups of the substrate, direct participation of a potassium ion in enzyme catalysis is strongly suggested.",
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AU - Masuda, Jun

AU - Tobimatsu, Takamasa

AU - Toraya, Tetsuo

AU - Suto, Kyoko

AU - Morimoto, Yukio

AU - Yasuoka, Noritake

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N2 - Background: Diol dehydratase is an enzyme that catalyzes the adenosylcobalamin (coenzyme B12) dependent conversion of 1,2-diols to the corresponding aldehydes. The reaction initiated by homolytic cleavage of the cobalt-carbon bond of the coenzyme proceeds by a radical mechanism. The enzyme is an (α2β2γ2 heterooligomer and has an absolute requirement for a potassium ion for catalytic activity. The crystal structure analysis of a diol dehydratase-cyanocobalamin complex was carried out in order to help understand the mechanism of action of this enzyme. Results: The three- dimensional structure of diol dehydratase in complex with cyanocobalamin was determined at 2.2 Å, resolution. The enzyme exists as a dimer of heterotrimers (αβγ)2. The cobalamin molecule is bound between the α and β subunits in the 'base-on' mode, that is, 5,6-dimethylbenzimidazole of the nucleotide moiety coordinates to the cobalt atom in the lower axial position. The α subunit includes a (β/α)8 barrel. The substrate, 1,2-propanediol, and an essential potassium ion are deeply buried inside the barrel. The two hydroxyl groups of the substrate coordinate directly to the potassium ion. Conclusions: This is the first crystallographic indication of the 'base-on' mode of cobalamin binding. An unusually long cobalt-base bond seems to favor homolytic cleavage of the cobalt-carbon bond and therefore to favor radical enzyme catalysis. Reactive radical intermediates can be protected from side reactions by spatial isolation inside the barrel. On the basis of unique direct interactions between the potassium ion and the two hydroxyl groups of the substrate, direct participation of a potassium ion in enzyme catalysis is strongly suggested.

AB - Background: Diol dehydratase is an enzyme that catalyzes the adenosylcobalamin (coenzyme B12) dependent conversion of 1,2-diols to the corresponding aldehydes. The reaction initiated by homolytic cleavage of the cobalt-carbon bond of the coenzyme proceeds by a radical mechanism. The enzyme is an (α2β2γ2 heterooligomer and has an absolute requirement for a potassium ion for catalytic activity. The crystal structure analysis of a diol dehydratase-cyanocobalamin complex was carried out in order to help understand the mechanism of action of this enzyme. Results: The three- dimensional structure of diol dehydratase in complex with cyanocobalamin was determined at 2.2 Å, resolution. The enzyme exists as a dimer of heterotrimers (αβγ)2. The cobalamin molecule is bound between the α and β subunits in the 'base-on' mode, that is, 5,6-dimethylbenzimidazole of the nucleotide moiety coordinates to the cobalt atom in the lower axial position. The α subunit includes a (β/α)8 barrel. The substrate, 1,2-propanediol, and an essential potassium ion are deeply buried inside the barrel. The two hydroxyl groups of the substrate coordinate directly to the potassium ion. Conclusions: This is the first crystallographic indication of the 'base-on' mode of cobalamin binding. An unusually long cobalt-base bond seems to favor homolytic cleavage of the cobalt-carbon bond and therefore to favor radical enzyme catalysis. Reactive radical intermediates can be protected from side reactions by spatial isolation inside the barrel. On the basis of unique direct interactions between the potassium ion and the two hydroxyl groups of the substrate, direct participation of a potassium ion in enzyme catalysis is strongly suggested.

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