1H NMR study of the dynamics of the pH modulation of axial coordination in Aplysia limacina (Val(E7)) and sperm whale double mutant His(E7) → Val,Thr(E10) → Arg metmyoglobin: Role of distal hydrogen bonding in ligation

Jun Qin, Usha Pande, Gerd N. La Mar, Franca Ascoli, Paolo Ascenzi, Francesca Cutruzzolá, Carlo Travaglini-Allocatelli, Maurizio Brunori

Research output: Contribution to journalArticle

Abstract

The molecular and electronic structure, thermodynamics, dynamics, and mechanism of interconversion of the pH-modulated reversible equilibria of Aplysia limacina metmyoglobin, (metMb), have been investigated by 1H NMR spectroscopy. The four identified species which interconvert slowly on the NMR time scale (lifetime > 1 ms) are metMbOH (B) at alkaline pH, five coordinate metMb (N) at acidic to neutral pH, an acidic form, A, near pH ∼4 and an extremely low pH form, D, attributed to an equilibrium unfolded species. The presence of strong distal hydrogen bonding by Arg (E10) to bound hydroxide is detected via a significant solvent isotope effect on the metMbOH (B) hyperfine shifts. Integration of the peak intensities yields pK values of 7.7 and ∼4 for the B ⇄ N and N ⇄ A equilibria, respectively. Saturation transfer via chemical exchange is observed for B ⇄ N and N ⇄ A, where the rates for forming metMbOH (B) and the acidic form A from N are base- and acid-catalyzed, respectively, while the reverse rates are first-order. The much slower interconversion rate for N ⇄ B in A. limacina metMb than His(E7) containing mammalian metMb is attributed to the fact that a ligand bond is broken rather than just proton transferred and that the equilibrium involves a major rearrangement of the orientation of Arg(E10). This conclusion is supported by 1H NMR data for the sperm whale double mutant His(E7) → Val/Thr(E10) → Arg metMb, which exhibits a pK ∼8.7 for the equilibrium between five-coordinate metMb (N) and metMbOH (B) with an even slower interconversion rate than in A. limacina metMb. This double mutant metMbOH (B) exhibits hydrogen bonding by Arg (E10) with coordinated hydroxide similar to that in A. limacina metMbOH. The slow but acid-catalyzed rates of conversion of A. limacina metMb (N) to the acid species A with significantly weakened bonding of the heme iron to the axial His(F8) residue is consistent with protonation of an inaccessible residue and/or a structural change accompanying the protonation equilibrium. It is concluded that metMb will coordinate water strongly only when there is a distal hydrogen bond acceptor residue, while the hydroxide ion is coordinated strongly only if there is a distal hydrogen bond donor residue.

Original languageEnglish
Pages (from-to)24012-24021
Number of pages10
JournalJournal of Biological Chemistry
Volume268
Issue number32
Publication statusPublished - Nov 15 1993

Fingerprint

Sperm Whale
Metmyoglobin
Aplysia
Hydrogen Bonding
Ligation
Hydrogen bonds
Nuclear magnetic resonance
Modulation
Protonation
Acids
Hydrogen
Proton Magnetic Resonance Spectroscopy
Molecular Structure
Heme
Thermodynamics
Isotopes
Molecular structure
Nuclear magnetic resonance spectroscopy
Electronic structure
Protons

ASJC Scopus subject areas

  • Biochemistry

Cite this

1H NMR study of the dynamics of the pH modulation of axial coordination in Aplysia limacina (Val(E7)) and sperm whale double mutant His(E7) → Val,Thr(E10) → Arg metmyoglobin : Role of distal hydrogen bonding in ligation. / Qin, Jun; Pande, Usha; La Mar, Gerd N.; Ascoli, Franca; Ascenzi, Paolo; Cutruzzolá, Francesca; Travaglini-Allocatelli, Carlo; Brunori, Maurizio.

In: Journal of Biological Chemistry, Vol. 268, No. 32, 15.11.1993, p. 24012-24021.

Research output: Contribution to journalArticle

Qin, Jun ; Pande, Usha ; La Mar, Gerd N. ; Ascoli, Franca ; Ascenzi, Paolo ; Cutruzzolá, Francesca ; Travaglini-Allocatelli, Carlo ; Brunori, Maurizio. / 1H NMR study of the dynamics of the pH modulation of axial coordination in Aplysia limacina (Val(E7)) and sperm whale double mutant His(E7) → Val,Thr(E10) → Arg metmyoglobin : Role of distal hydrogen bonding in ligation. In: Journal of Biological Chemistry. 1993 ; Vol. 268, No. 32. pp. 24012-24021.
@article{fbcc15d00b6143cf9f6d38b265ebd1b9,
title = "1H NMR study of the dynamics of the pH modulation of axial coordination in Aplysia limacina (Val(E7)) and sperm whale double mutant His(E7) → Val,Thr(E10) → Arg metmyoglobin: Role of distal hydrogen bonding in ligation",
abstract = "The molecular and electronic structure, thermodynamics, dynamics, and mechanism of interconversion of the pH-modulated reversible equilibria of Aplysia limacina metmyoglobin, (metMb), have been investigated by 1H NMR spectroscopy. The four identified species which interconvert slowly on the NMR time scale (lifetime > 1 ms) are metMbOH (B) at alkaline pH, five coordinate metMb (N) at acidic to neutral pH, an acidic form, A, near pH ∼4 and an extremely low pH form, D, attributed to an equilibrium unfolded species. The presence of strong distal hydrogen bonding by Arg (E10) to bound hydroxide is detected via a significant solvent isotope effect on the metMbOH (B) hyperfine shifts. Integration of the peak intensities yields pK values of 7.7 and ∼4 for the B ⇄ N and N ⇄ A equilibria, respectively. Saturation transfer via chemical exchange is observed for B ⇄ N and N ⇄ A, where the rates for forming metMbOH (B) and the acidic form A from N are base- and acid-catalyzed, respectively, while the reverse rates are first-order. The much slower interconversion rate for N ⇄ B in A. limacina metMb than His(E7) containing mammalian metMb is attributed to the fact that a ligand bond is broken rather than just proton transferred and that the equilibrium involves a major rearrangement of the orientation of Arg(E10). This conclusion is supported by 1H NMR data for the sperm whale double mutant His(E7) → Val/Thr(E10) → Arg metMb, which exhibits a pK ∼8.7 for the equilibrium between five-coordinate metMb (N) and metMbOH (B) with an even slower interconversion rate than in A. limacina metMb. This double mutant metMbOH (B) exhibits hydrogen bonding by Arg (E10) with coordinated hydroxide similar to that in A. limacina metMbOH. The slow but acid-catalyzed rates of conversion of A. limacina metMb (N) to the acid species A with significantly weakened bonding of the heme iron to the axial His(F8) residue is consistent with protonation of an inaccessible residue and/or a structural change accompanying the protonation equilibrium. It is concluded that metMb will coordinate water strongly only when there is a distal hydrogen bond acceptor residue, while the hydroxide ion is coordinated strongly only if there is a distal hydrogen bond donor residue.",
author = "Jun Qin and Usha Pande and {La Mar}, {Gerd N.} and Franca Ascoli and Paolo Ascenzi and Francesca Cutruzzol{\'a} and Carlo Travaglini-Allocatelli and Maurizio Brunori",
year = "1993",
month = "11",
day = "15",
language = "English",
volume = "268",
pages = "24012--24021",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "32",

}

TY - JOUR

T1 - 1H NMR study of the dynamics of the pH modulation of axial coordination in Aplysia limacina (Val(E7)) and sperm whale double mutant His(E7) → Val,Thr(E10) → Arg metmyoglobin

T2 - Role of distal hydrogen bonding in ligation

AU - Qin, Jun

AU - Pande, Usha

AU - La Mar, Gerd N.

AU - Ascoli, Franca

AU - Ascenzi, Paolo

AU - Cutruzzolá, Francesca

AU - Travaglini-Allocatelli, Carlo

AU - Brunori, Maurizio

PY - 1993/11/15

Y1 - 1993/11/15

N2 - The molecular and electronic structure, thermodynamics, dynamics, and mechanism of interconversion of the pH-modulated reversible equilibria of Aplysia limacina metmyoglobin, (metMb), have been investigated by 1H NMR spectroscopy. The four identified species which interconvert slowly on the NMR time scale (lifetime > 1 ms) are metMbOH (B) at alkaline pH, five coordinate metMb (N) at acidic to neutral pH, an acidic form, A, near pH ∼4 and an extremely low pH form, D, attributed to an equilibrium unfolded species. The presence of strong distal hydrogen bonding by Arg (E10) to bound hydroxide is detected via a significant solvent isotope effect on the metMbOH (B) hyperfine shifts. Integration of the peak intensities yields pK values of 7.7 and ∼4 for the B ⇄ N and N ⇄ A equilibria, respectively. Saturation transfer via chemical exchange is observed for B ⇄ N and N ⇄ A, where the rates for forming metMbOH (B) and the acidic form A from N are base- and acid-catalyzed, respectively, while the reverse rates are first-order. The much slower interconversion rate for N ⇄ B in A. limacina metMb than His(E7) containing mammalian metMb is attributed to the fact that a ligand bond is broken rather than just proton transferred and that the equilibrium involves a major rearrangement of the orientation of Arg(E10). This conclusion is supported by 1H NMR data for the sperm whale double mutant His(E7) → Val/Thr(E10) → Arg metMb, which exhibits a pK ∼8.7 for the equilibrium between five-coordinate metMb (N) and metMbOH (B) with an even slower interconversion rate than in A. limacina metMb. This double mutant metMbOH (B) exhibits hydrogen bonding by Arg (E10) with coordinated hydroxide similar to that in A. limacina metMbOH. The slow but acid-catalyzed rates of conversion of A. limacina metMb (N) to the acid species A with significantly weakened bonding of the heme iron to the axial His(F8) residue is consistent with protonation of an inaccessible residue and/or a structural change accompanying the protonation equilibrium. It is concluded that metMb will coordinate water strongly only when there is a distal hydrogen bond acceptor residue, while the hydroxide ion is coordinated strongly only if there is a distal hydrogen bond donor residue.

AB - The molecular and electronic structure, thermodynamics, dynamics, and mechanism of interconversion of the pH-modulated reversible equilibria of Aplysia limacina metmyoglobin, (metMb), have been investigated by 1H NMR spectroscopy. The four identified species which interconvert slowly on the NMR time scale (lifetime > 1 ms) are metMbOH (B) at alkaline pH, five coordinate metMb (N) at acidic to neutral pH, an acidic form, A, near pH ∼4 and an extremely low pH form, D, attributed to an equilibrium unfolded species. The presence of strong distal hydrogen bonding by Arg (E10) to bound hydroxide is detected via a significant solvent isotope effect on the metMbOH (B) hyperfine shifts. Integration of the peak intensities yields pK values of 7.7 and ∼4 for the B ⇄ N and N ⇄ A equilibria, respectively. Saturation transfer via chemical exchange is observed for B ⇄ N and N ⇄ A, where the rates for forming metMbOH (B) and the acidic form A from N are base- and acid-catalyzed, respectively, while the reverse rates are first-order. The much slower interconversion rate for N ⇄ B in A. limacina metMb than His(E7) containing mammalian metMb is attributed to the fact that a ligand bond is broken rather than just proton transferred and that the equilibrium involves a major rearrangement of the orientation of Arg(E10). This conclusion is supported by 1H NMR data for the sperm whale double mutant His(E7) → Val/Thr(E10) → Arg metMb, which exhibits a pK ∼8.7 for the equilibrium between five-coordinate metMb (N) and metMbOH (B) with an even slower interconversion rate than in A. limacina metMb. This double mutant metMbOH (B) exhibits hydrogen bonding by Arg (E10) with coordinated hydroxide similar to that in A. limacina metMbOH. The slow but acid-catalyzed rates of conversion of A. limacina metMb (N) to the acid species A with significantly weakened bonding of the heme iron to the axial His(F8) residue is consistent with protonation of an inaccessible residue and/or a structural change accompanying the protonation equilibrium. It is concluded that metMb will coordinate water strongly only when there is a distal hydrogen bond acceptor residue, while the hydroxide ion is coordinated strongly only if there is a distal hydrogen bond donor residue.

UR - http://www.scopus.com/inward/record.url?scp=0027918370&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0027918370&partnerID=8YFLogxK

M3 - Article

C2 - 8226945

AN - SCOPUS:0027918370

VL - 268

SP - 24012

EP - 24021

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 32

ER -