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Direct visualization of critical hydrogen atoms in a pyridoxal 5'-phosphate enzyme

Journal Article · · Nature Communications
 [1];  [2];  [2];  [3];  [4];  [2];  [5];  [2];  [6]
  1. Univ. of Toledo, Toledo, OH (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
  2. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
  3. Institut Laue Langevin, Grenoble Cedex (France)
  4. Rutherford Appleton Lab., Didcot (United Kingdom)
  5. Univ. of Tennessee, Knoxville, TN (United States)
  6. Univ. of Toledo, Toledo, OH (United States)
+ Show Author Affiliations
Enzymes dependent on pyridoxal 5'-phosphate (PLP, the active form of vitamin B6) perform a myriad of diverse chemical transformations. They promote various reactions by modulating the electronic states of PLP through weak interactions in the active site. Neutron crystallography has the unique ability of visualizing the nuclear positions of hydrogen atoms in macromolecules. Here we present a room-temperature neutron structure of a homodimeric PLP-dependent enzyme, aspartate aminotransferase, which was reacted in situ with α-methylaspartate. In one monomer, the PLP remained as an internal aldimine with a deprotonated Schiff base. In the second monomer, the external aldimine formed with the substrate analog. We observe a deuterium equidistant between the Schiff base and the C-terminal carboxylate of the substrate, a position indicative of a low-barrier hydrogen bond. As a result, quantum chemical calculations and a low-pH room-temperature X-ray structure provide insight into the physical phenomena that control the electronic modulation in aspartate aminotransferase.
Research Organization:
Oak Ridge National Laboratory (ORNL), Oak Ridge, TN (United States)
Sponsoring Organization:
USDOE
Grant/Contract Number:
AC05-00OR22725
OSTI ID:
1407771
Journal Information:
Nature Communications, Journal Name: Nature Communications Journal Issue: 1 Vol. 8; ISSN 2041-1723
Publisher:
Nature Publishing GroupCopyright Statement
Country of Publication:
United States
Language:
English

References (54)

Long-Range Electrostatics-Induced Two-Proton Transfer Captured by Neutron Crystallography in an Enzyme Catalytic Site journal March 2016
NBO 6.0 : Natural bond orbital analysis program journal March 2013
Hyperconjugation journal January 2011
Catalytic isomerization of glucose to fructose over organic ligands: a DFT study journal December 2022
Investigations on the E/Z-isomerism of neonicotinoids journal July 2020
Reaction specificity in pyridoxal phosphate enzymes journal January 2005
Aspartate aminotransferase: An old dog teaches new tricks journal February 2014
Lysine 258 in aspartate aminotransferase: Enforcer of the Circe effect for amino acid substrates and the general-base catalyst for the 1,3-prototropic shift journal February 1993
Use of 1 H− 15 N Heteronuclear Multiple-Quantum Coherence NMR Spectroscopy To Study the Active Site of Aspartate Aminotransferase † journal January 1997
Transient-State Kinetics of the Reaction of Aspartate Aminotransferase with Aspartate at Low pH Reveals Dual Routes in the Enzyme−Substrate Association Process † journal November 1997
Glycine Enolates:  The Large Effect of Iminium Ion Formation on α-Amino Carbon Acidity journal August 2001
X-ray and NMR Crystallography in an Enzyme Active Site: The Indoline Quinonoid Intermediate in Tryptophan Synthase journal January 2011
NMR Studies of Protonation and Hydrogen Bond States of Internal Aldimines of Pyridoxal 5′-Phosphate Acid–Base in Alanine Racemase, Aspartate Aminotransferase, and Poly- l -lysine journal November 2013
Protonation States of the Tryptophan Synthase Internal Aldimine Active Site from Solid-State NMR Spectroscopy: Direct Observation of the Protonated Schiff Base Linkage to Pyridoxal-5′-Phosphate journal September 2014
NMR Crystallography of a Carbanionic Intermediate in Tryptophan Synthase: Chemical Structure, Tautomerization, and Reaction Specificity journal November 2016
15 N Nuclear Magnetic Resonance Studies of Acid−Base Properties of Pyridoxal-5‘-Phosphate Aldimines in Aqueous Solution journal April 2007
A Hybrid QM/MM Simulation Study of Intramolecular Proton Transfer in the Pyridoxal 5′-Phosphate in the Active Site of Transaminase: Influence of Active Site Interaction on Proton Transfer journal September 2014
A genomic overview of pyridoxal-phosphate-dependent enzymes journal September 2003
Theoretical studies of strong hydrogen bonds journal January 2006
Accurate Coulomb-fitting basis sets for H to Rn journal January 2006
Natural localized molecular orbitals journal August 1985
Conformation and reaction specificity in pyridoxal phosphate enzymes. journal April 1966
Refinement and Comparisons of the Crystal Structures of Pig Cytosolic Aspartate Aminotransferase and Its Complex with 2-Methylaspartate journal July 1997
Direct evidence that an extended hydrogen-bonding network influences activation of pyridoxal 5′-phosphate in aspartate aminotransferase journal February 2017
Conformational Change in Aspartate Aminotransferase on Substrate Binding Induces Strain in the Catalytic Group and Enhances Catalysis journal December 2002
LAUEGEN version 6.0 and INTLDM journal June 1998
Features and development of Coot journal March 2010
Neutron macromolecular crystallography with LADI-III journal October 2010
The IMAGINE instrument: first neutron protein structure and new capabilities for neutron macromolecular crystallography journal September 2013
Visualizing the Bohr effect in hemoglobin: neutron structure of equine cyanomethemoglobin in the R state and comparison with human deoxyhemoglobin in the T state journal June 2016
Low-barrier hydrogen bonds and enzymic catalysis journal June 1994
Neutron Diffraction in Studies of Protein Dynamics and Functions, Application of book January 2007
Resonance Raman spectra of the pyridoxal coenzyme in aspartate aminotransferase. Evidence for pyridine protonation and a novel photochemical H/D exchange at the imine carbon atom. journal September 1985
NMR studies of 1H resonances in the 10-18-ppm range for cytosolic aspartate aminotransferase. journal November 1994
NMR observation of exchangeable protons of pyridoxal phosphate and histidine residues in cytosolic aspartate aminotransferase. journal September 1991
[16] SHELXL: High-resolution refinement book January 1997
Critical hydrogen bonds and protonation states of pyridoxal 5′-phosphate revealed by NMR journal November 2011
Pyridoxal 5′-phosphate: electrophilic catalyst extraordinaire journal October 2009
Theoretical study on the distribution of atomic charges in the Schiff bases of 3-hydroxypyridine-4-aldehyde and alanine. The effect of the protonation state of the pyridine and imine nitrogen atoms journal January 2009
Universal Solvation Model Based on Solute Electron Density and on a Continuum Model of the Solvent Defined by the Bulk Dielectric Constant and Atomic Surface Tensions
  • Marenich, Aleksandr V.; Cramer, Christopher J.; Truhlar, Donald G.
  • The Journal of Physical Chemistry B, Vol. 113, Issue 18, p. 6378-6396 https://doi.org/10.1021/jp810292n
journal May 2009
Balanced basis sets of split valence, triple zeta valence and quadruple zeta valence quality for H to Rn: Design and assessment of accuracy journal January 2005
Direct determination of protonation states and visualization of hydrogen bonding in a glycoside hydrolase with neutron crystallography journal September 2015
Conformational Change in Aspartate Aminotransferase on Substrate Binding Induces Strain in the Catalytic Group and Enhances Catalysis journal December 2002
Global indicators of X-ray data quality journal April 2001
LAUEGEN , an X-windows-based program for the processing of Laue diffraction data journal April 1995
LAUEGEN version 6.0 and INTLDM journal June 1998
LSCALE – the new normalization, scaling and absorption correction program in the Daresbury Laue software suite journal June 1999
HKL -3000: the integration of data reduction and structure solution – from diffraction images to an initial model in minutes journal July 2006
Generalized X-ray and neutron crystallographic analysis: more accurate and complete structures for biological macromolecules journal May 2009
Features and development of Coot journal March 2010
Neutron macromolecular crystallography with LADI-III journal October 2010
The IMAGINE instrument: first neutron protein structure and new capabilities for neutron macromolecular crystallography journal September 2013
Visualizing the Bohr effect in hemoglobin: neutron structure of equine cyanomethemoglobin in the R state and comparison with human deoxyhemoglobin in the T state journal June 2016
Hyperconjugation in hydrocarbons: Not just a “mild sort of conjugation” journal April 2013

Cited By (7)

Dimerization misalignment in human glutamate-oxaloacetate transaminase variants is the primary factor for PLP release journal September 2018
Current Advances on Structure-Function Relationships of Pyridoxal 5′-Phosphate-Dependent Enzymes journal March 2019
Neutron Macromolecular Crystallography book January 2020
Hyperconjugation promoted by hydrogen bonding between His98/His241 and a carboxyl group contributes to tyrosine decarboxylase catalysis journal January 2019
Neutron macromolecular crystallography journal February 2018
Instrument and shielding design of a neutron diffractometer at J-PARC for protein crystallography covering crystals with large unit-cell volume journal April 2018
Neutron scattering in the biological sciences: progress and prospects journal December 2018

Figures / Tables (8)

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Table 1(p. 3)table Table 1
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Fig. 6(p. 6)figure Fig. 6
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