Hydrogen bonding interaction of the amide group of Asn and Gln at distal E7 of bovine myoglobin with bound-ligand and its functional consequences

Yasuhiko Yamamoto, Norimasa Kurihara, Tsuyoshi Egawa, Hideo Shimada, Yuzuru Ishimura

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Asn and Gln with an amide group at γ- and δ-positions, respectively, were substituted for distal His-E7 of bovine myoglobin to establish a system where hydrogen bonding interaction between the distal residue and bound-ligand can be altered by changing donor-acceptor distance. Two mutant myoglobins showed nearly identical 1H-NMR spectral pattern for resolved heme peripheral side-chain and amino acid proton signals and similar two-dimensional NMR connectivities irrespective of cyanide-bound and -unbound states, indicating that the heme electronic structure and the molecular structure of the active site are not affected by a difference in one methylene group at the E7 position. Chemical exchange rate of Asn-E7 N(δ)H proton in met-cyano myoglobin is larger than that of Gln-E7 N(ε)H proton by at least two orders of magnitude, suggesting a considerable difference in the strength of hydrogen bond between the E7 side-chain and bound-ligand, due to the differential donor-acceptor distance between the two mutants. Thus a comparative study between the two proteins provides an ideal system to delineate a relationship between the stabilization of bound-ligand by the hydrogen bond and myoglobin's ligand affinity. The Asn-mutant showed a faster dissociation of cyano ion from met-myoglobin than the Gln-mutant by over 30-fold. Similarly, oxygen dissociation is faster in the Asn-mutant than in the Gln-mutant by ~100-fold. Association of cyanide anion to the mutant met-myoglobin was accelerated by changing Gln to Asn by a 4-fold. Likewise, oxygen binding was accelerated by ~2-fold by the above substitution. The present findings confirm that hydrogen bonding with the distal residue is a dominant factor for determining the ligand dissociation rate, whereas steric hindrance exerted by the distal residue is a primary determinant for the ligand association. Copyright (C) 1999 Elsevier Science B.V.

Original languageEnglish (US)
Pages (from-to)27-44
Number of pages18
JournalBiochimica et Biophysica Acta - Protein Structure and Molecular Enzymology
Volume1433
Issue number1-2
DOIs
StatePublished - Aug 17 1999
Externally publishedYes

Fingerprint

Myoglobin
Hydrogen Bonding
Amides
Hydrogen bonds
Ligands
Metmyoglobin
Protons
Cyanides
Heme
Hydrogen
Nuclear magnetic resonance
Oxygen
Molecular Structure
Molecular structure
Electronic structure
Anions
Catalytic Domain
Substitution reactions
Stabilization
Association reactions

Keywords

  • Hydrogen bond
  • Ligand affinity
  • Myoglobin mutant
  • NMR
  • Paramagnetic relaxation
  • Paramagnetic shift

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Structural Biology
  • Biophysics

Cite this

Hydrogen bonding interaction of the amide group of Asn and Gln at distal E7 of bovine myoglobin with bound-ligand and its functional consequences. / Yamamoto, Yasuhiko; Kurihara, Norimasa; Egawa, Tsuyoshi; Shimada, Hideo; Ishimura, Yuzuru.

In: Biochimica et Biophysica Acta - Protein Structure and Molecular Enzymology, Vol. 1433, No. 1-2, 17.08.1999, p. 27-44.

Research output: Contribution to journalArticle

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AU - Shimada, Hideo

AU - Ishimura, Yuzuru

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N2 - Asn and Gln with an amide group at γ- and δ-positions, respectively, were substituted for distal His-E7 of bovine myoglobin to establish a system where hydrogen bonding interaction between the distal residue and bound-ligand can be altered by changing donor-acceptor distance. Two mutant myoglobins showed nearly identical 1H-NMR spectral pattern for resolved heme peripheral side-chain and amino acid proton signals and similar two-dimensional NMR connectivities irrespective of cyanide-bound and -unbound states, indicating that the heme electronic structure and the molecular structure of the active site are not affected by a difference in one methylene group at the E7 position. Chemical exchange rate of Asn-E7 N(δ)H proton in met-cyano myoglobin is larger than that of Gln-E7 N(ε)H proton by at least two orders of magnitude, suggesting a considerable difference in the strength of hydrogen bond between the E7 side-chain and bound-ligand, due to the differential donor-acceptor distance between the two mutants. Thus a comparative study between the two proteins provides an ideal system to delineate a relationship between the stabilization of bound-ligand by the hydrogen bond and myoglobin's ligand affinity. The Asn-mutant showed a faster dissociation of cyano ion from met-myoglobin than the Gln-mutant by over 30-fold. Similarly, oxygen dissociation is faster in the Asn-mutant than in the Gln-mutant by ~100-fold. Association of cyanide anion to the mutant met-myoglobin was accelerated by changing Gln to Asn by a 4-fold. Likewise, oxygen binding was accelerated by ~2-fold by the above substitution. The present findings confirm that hydrogen bonding with the distal residue is a dominant factor for determining the ligand dissociation rate, whereas steric hindrance exerted by the distal residue is a primary determinant for the ligand association. Copyright (C) 1999 Elsevier Science B.V.

AB - Asn and Gln with an amide group at γ- and δ-positions, respectively, were substituted for distal His-E7 of bovine myoglobin to establish a system where hydrogen bonding interaction between the distal residue and bound-ligand can be altered by changing donor-acceptor distance. Two mutant myoglobins showed nearly identical 1H-NMR spectral pattern for resolved heme peripheral side-chain and amino acid proton signals and similar two-dimensional NMR connectivities irrespective of cyanide-bound and -unbound states, indicating that the heme electronic structure and the molecular structure of the active site are not affected by a difference in one methylene group at the E7 position. Chemical exchange rate of Asn-E7 N(δ)H proton in met-cyano myoglobin is larger than that of Gln-E7 N(ε)H proton by at least two orders of magnitude, suggesting a considerable difference in the strength of hydrogen bond between the E7 side-chain and bound-ligand, due to the differential donor-acceptor distance between the two mutants. Thus a comparative study between the two proteins provides an ideal system to delineate a relationship between the stabilization of bound-ligand by the hydrogen bond and myoglobin's ligand affinity. The Asn-mutant showed a faster dissociation of cyano ion from met-myoglobin than the Gln-mutant by over 30-fold. Similarly, oxygen dissociation is faster in the Asn-mutant than in the Gln-mutant by ~100-fold. Association of cyanide anion to the mutant met-myoglobin was accelerated by changing Gln to Asn by a 4-fold. Likewise, oxygen binding was accelerated by ~2-fold by the above substitution. The present findings confirm that hydrogen bonding with the distal residue is a dominant factor for determining the ligand dissociation rate, whereas steric hindrance exerted by the distal residue is a primary determinant for the ligand association. Copyright (C) 1999 Elsevier Science B.V.

KW - Hydrogen bond

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KW - Myoglobin mutant

KW - NMR

KW - Paramagnetic relaxation

KW - Paramagnetic shift

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