Evolution of enzymatic activities in the enolase superfamily

D-mannonate dehydratase from Novosphingobium aromaticivorans

John F. Rakus, Alexander A. Fedorov, Elena V. Fedorov, Margaret E. Glasner, Jacob E. Vick, Patricia C. Babbitt, Steven C. Almo, John A. Gerlt

Research output: Contribution to journalArticle

30 Citations (Scopus)

Abstract

The D-mannonate dehydratase (ManD) function was assigned to a group of orthologous proteins in the mechanistically diverse enolase superfamily by screening a library of acid sugars. Structures of the wild type ManD from Novosphingobium aromaticivorans were determined at pH 7.5 in the presence of Mg2+ and also in the presence of Mg2+ and the 2-keto-3-keto-D-gluconate dehydration product; the structure of the catalytically active K271E mutant was determined at pH 5.5 in the presence of the D-mannonate substrate. As previously observed in the structures of other members of the enolase superfamily, ManD contains two domains, an N-terminal α+β capping domain and a (β/α)7β-barrel domain. The barrel domain contains the ligands for the essential Mg 2+, Asp 210, Glu 236, and Glu 262, at the ends of the third, fourth, and fifth β-strands of the barrel domain, respectively. However, the barrel domain lacks both the Lys acid/base catalyst at the end of the second β-strand and the His-Asp dyad acid/base catalyst at the ends of the seventh and sixth β-strands, respectively, that are found in many members of the superfamily. Instead, a hydrogen-bonded dyad of Tyr 159 in a loop following the second β-strand and Arg 147 at the end of the second β-strand are positioned to initiate the reaction by abstraction of the 2-proton. Both Tyr 159 and His 212, at the end of the third β-strand, are positioned to facilitate both syn-dehydration and ketonization of the resulting enol intermediate to yield the 2-keto-3-keto-D-gluconate product with the observed retention of configuration. The identities and locations of these acid/base catalysts as well as of cationic amino acid residues that stabilize the enolate anion intermediate define a new structural strategy for catalysis (subgroup) in the mechanistically diverse enolase superfamily. With these differences, we provide additional evidence that the ligands for the essential Mg2+ are the only conserved residues in the enolase superfamily, establishing the primary functional importance of the Mg2+-assisted strategy for stabilizing the enolate anion intermediate.

Original languageEnglish (US)
Pages (from-to)12896-12908
Number of pages13
JournalBiochemistry
Volume46
Issue number45
DOIs
StatePublished - Nov 13 2007

Fingerprint

Phosphopyruvate Hydratase
Dehydration
Catalysts
Acids
Anions
Ligands
Sugar Acids
Catalysis
Protons
Hydrogen
Screening
Amino Acids
mannonate dehydratase
Substrates
Proteins
gluconic acid

ASJC Scopus subject areas

  • Biochemistry

Cite this

Rakus, J. F., Fedorov, A. A., Fedorov, E. V., Glasner, M. E., Vick, J. E., Babbitt, P. C., ... Gerlt, J. A. (2007). Evolution of enzymatic activities in the enolase superfamily: D-mannonate dehydratase from Novosphingobium aromaticivorans. Biochemistry, 46(45), 12896-12908. https://doi.org/10.1021/bi701703w

Evolution of enzymatic activities in the enolase superfamily : D-mannonate dehydratase from Novosphingobium aromaticivorans. / Rakus, John F.; Fedorov, Alexander A.; Fedorov, Elena V.; Glasner, Margaret E.; Vick, Jacob E.; Babbitt, Patricia C.; Almo, Steven C.; Gerlt, John A.

In: Biochemistry, Vol. 46, No. 45, 13.11.2007, p. 12896-12908.

Research output: Contribution to journalArticle

Rakus, JF, Fedorov, AA, Fedorov, EV, Glasner, ME, Vick, JE, Babbitt, PC, Almo, SC & Gerlt, JA 2007, 'Evolution of enzymatic activities in the enolase superfamily: D-mannonate dehydratase from Novosphingobium aromaticivorans', Biochemistry, vol. 46, no. 45, pp. 12896-12908. https://doi.org/10.1021/bi701703w
Rakus, John F. ; Fedorov, Alexander A. ; Fedorov, Elena V. ; Glasner, Margaret E. ; Vick, Jacob E. ; Babbitt, Patricia C. ; Almo, Steven C. ; Gerlt, John A. / Evolution of enzymatic activities in the enolase superfamily : D-mannonate dehydratase from Novosphingobium aromaticivorans. In: Biochemistry. 2007 ; Vol. 46, No. 45. pp. 12896-12908.
@article{f13d237d18f94679a29242c09b6ff009,
title = "Evolution of enzymatic activities in the enolase superfamily: D-mannonate dehydratase from Novosphingobium aromaticivorans",
abstract = "The D-mannonate dehydratase (ManD) function was assigned to a group of orthologous proteins in the mechanistically diverse enolase superfamily by screening a library of acid sugars. Structures of the wild type ManD from Novosphingobium aromaticivorans were determined at pH 7.5 in the presence of Mg2+ and also in the presence of Mg2+ and the 2-keto-3-keto-D-gluconate dehydration product; the structure of the catalytically active K271E mutant was determined at pH 5.5 in the presence of the D-mannonate substrate. As previously observed in the structures of other members of the enolase superfamily, ManD contains two domains, an N-terminal α+β capping domain and a (β/α)7β-barrel domain. The barrel domain contains the ligands for the essential Mg 2+, Asp 210, Glu 236, and Glu 262, at the ends of the third, fourth, and fifth β-strands of the barrel domain, respectively. However, the barrel domain lacks both the Lys acid/base catalyst at the end of the second β-strand and the His-Asp dyad acid/base catalyst at the ends of the seventh and sixth β-strands, respectively, that are found in many members of the superfamily. Instead, a hydrogen-bonded dyad of Tyr 159 in a loop following the second β-strand and Arg 147 at the end of the second β-strand are positioned to initiate the reaction by abstraction of the 2-proton. Both Tyr 159 and His 212, at the end of the third β-strand, are positioned to facilitate both syn-dehydration and ketonization of the resulting enol intermediate to yield the 2-keto-3-keto-D-gluconate product with the observed retention of configuration. The identities and locations of these acid/base catalysts as well as of cationic amino acid residues that stabilize the enolate anion intermediate define a new structural strategy for catalysis (subgroup) in the mechanistically diverse enolase superfamily. With these differences, we provide additional evidence that the ligands for the essential Mg2+ are the only conserved residues in the enolase superfamily, establishing the primary functional importance of the Mg2+-assisted strategy for stabilizing the enolate anion intermediate.",
author = "Rakus, {John F.} and Fedorov, {Alexander A.} and Fedorov, {Elena V.} and Glasner, {Margaret E.} and Vick, {Jacob E.} and Babbitt, {Patricia C.} and Almo, {Steven C.} and Gerlt, {John A.}",
year = "2007",
month = "11",
day = "13",
doi = "10.1021/bi701703w",
language = "English (US)",
volume = "46",
pages = "12896--12908",
journal = "Biochemistry",
issn = "0006-2960",
publisher = "American Chemical Society",
number = "45",

}

TY - JOUR

T1 - Evolution of enzymatic activities in the enolase superfamily

T2 - D-mannonate dehydratase from Novosphingobium aromaticivorans

AU - Rakus, John F.

AU - Fedorov, Alexander A.

AU - Fedorov, Elena V.

AU - Glasner, Margaret E.

AU - Vick, Jacob E.

AU - Babbitt, Patricia C.

AU - Almo, Steven C.

AU - Gerlt, John A.

PY - 2007/11/13

Y1 - 2007/11/13

N2 - The D-mannonate dehydratase (ManD) function was assigned to a group of orthologous proteins in the mechanistically diverse enolase superfamily by screening a library of acid sugars. Structures of the wild type ManD from Novosphingobium aromaticivorans were determined at pH 7.5 in the presence of Mg2+ and also in the presence of Mg2+ and the 2-keto-3-keto-D-gluconate dehydration product; the structure of the catalytically active K271E mutant was determined at pH 5.5 in the presence of the D-mannonate substrate. As previously observed in the structures of other members of the enolase superfamily, ManD contains two domains, an N-terminal α+β capping domain and a (β/α)7β-barrel domain. The barrel domain contains the ligands for the essential Mg 2+, Asp 210, Glu 236, and Glu 262, at the ends of the third, fourth, and fifth β-strands of the barrel domain, respectively. However, the barrel domain lacks both the Lys acid/base catalyst at the end of the second β-strand and the His-Asp dyad acid/base catalyst at the ends of the seventh and sixth β-strands, respectively, that are found in many members of the superfamily. Instead, a hydrogen-bonded dyad of Tyr 159 in a loop following the second β-strand and Arg 147 at the end of the second β-strand are positioned to initiate the reaction by abstraction of the 2-proton. Both Tyr 159 and His 212, at the end of the third β-strand, are positioned to facilitate both syn-dehydration and ketonization of the resulting enol intermediate to yield the 2-keto-3-keto-D-gluconate product with the observed retention of configuration. The identities and locations of these acid/base catalysts as well as of cationic amino acid residues that stabilize the enolate anion intermediate define a new structural strategy for catalysis (subgroup) in the mechanistically diverse enolase superfamily. With these differences, we provide additional evidence that the ligands for the essential Mg2+ are the only conserved residues in the enolase superfamily, establishing the primary functional importance of the Mg2+-assisted strategy for stabilizing the enolate anion intermediate.

AB - The D-mannonate dehydratase (ManD) function was assigned to a group of orthologous proteins in the mechanistically diverse enolase superfamily by screening a library of acid sugars. Structures of the wild type ManD from Novosphingobium aromaticivorans were determined at pH 7.5 in the presence of Mg2+ and also in the presence of Mg2+ and the 2-keto-3-keto-D-gluconate dehydration product; the structure of the catalytically active K271E mutant was determined at pH 5.5 in the presence of the D-mannonate substrate. As previously observed in the structures of other members of the enolase superfamily, ManD contains two domains, an N-terminal α+β capping domain and a (β/α)7β-barrel domain. The barrel domain contains the ligands for the essential Mg 2+, Asp 210, Glu 236, and Glu 262, at the ends of the third, fourth, and fifth β-strands of the barrel domain, respectively. However, the barrel domain lacks both the Lys acid/base catalyst at the end of the second β-strand and the His-Asp dyad acid/base catalyst at the ends of the seventh and sixth β-strands, respectively, that are found in many members of the superfamily. Instead, a hydrogen-bonded dyad of Tyr 159 in a loop following the second β-strand and Arg 147 at the end of the second β-strand are positioned to initiate the reaction by abstraction of the 2-proton. Both Tyr 159 and His 212, at the end of the third β-strand, are positioned to facilitate both syn-dehydration and ketonization of the resulting enol intermediate to yield the 2-keto-3-keto-D-gluconate product with the observed retention of configuration. The identities and locations of these acid/base catalysts as well as of cationic amino acid residues that stabilize the enolate anion intermediate define a new structural strategy for catalysis (subgroup) in the mechanistically diverse enolase superfamily. With these differences, we provide additional evidence that the ligands for the essential Mg2+ are the only conserved residues in the enolase superfamily, establishing the primary functional importance of the Mg2+-assisted strategy for stabilizing the enolate anion intermediate.

UR - http://www.scopus.com/inward/record.url?scp=36049048325&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=36049048325&partnerID=8YFLogxK

U2 - 10.1021/bi701703w

DO - 10.1021/bi701703w

M3 - Article

VL - 46

SP - 12896

EP - 12908

JO - Biochemistry

JF - Biochemistry

SN - 0006-2960

IS - 45

ER -