Nitrogen-15 NMR spectroscopy of hydrogen-bonding interactions in the active site of serine proteases: evidence for a moving histidine mechanism

Biochemistry ◽  
1986 ◽  
Vol 25 (23) ◽  
pp. 7751-7759 ◽  
Author(s):  
William W. Bachovchin
Keyword(s):  
Hydrogen Bonding ◽  
Nmr Spectroscopy ◽  
Active Site ◽  
Serine Proteases ◽  
Hydrogen Bonding Interactions

scholarly journals The Geometry of the Catalytic Active Site in [FeFe]-Hydrogenases is Determined by Hydrogen Bonding and Proton Transfer

2019 ◽  
Author(s):  
Jifu Duan ◽  
Stefan Mebs ◽  
Moritz Senger ◽  
Konstantin Laun ◽  
Florian Wittkamp ◽  
...  
Keyword(s):  
Hydrogen Bonding ◽  
Proton Transfer ◽  
Active Site ◽  
Time Resolved ◽  
X Ray Crystallography ◽  
Hydrogen Bonding Interactions ◽  
Catalytic Active Site ◽  
Catalytic Intermediates ◽  
Time Resolved Infrared Spectroscopy

The H2 conversion and CO inhibition reactivity of nine [FeFe]-hydrogenase constructs with semi-artificial cofactors was studied by in situ and time-resolved infrared spectroscopy, X-ray crystallography, and theoretical methods. Impaired hydrogen turnover and proton transfer as well as characteristic CO inhibition/ reactivation kinetics are assigned to varying degrees of hydrogen-bonding interactions at the active site. We show that the probability to adopt catalytic intermediates is modulated by intramolecular and protein-cofactor interactions that govern structural dynamics at the active site of [FeFe]-hydrogenases.<br>

scholarly journals A crystallographic study of the Toho-1 β-lactamase acylation mechanism

2014 ◽  
Vol 70 (a1) ◽  
pp. C1207-C1207
Author(s):  
Leighton Coates
Keyword(s):  
Hydrogen Bonding ◽  
Transition State ◽  
Active Site ◽  
Catalytic Mechanism ◽  
Substrate Binding ◽  
Ligand Complex ◽  
Transition State Analog ◽  
Hydrogen Bonding Interactions ◽  
Active Site Region ◽  
Neutron Crystallography

β-lactam antibiotics have been used effectively over several decades against many types of highly virulent bacteria. The predominant cause of resistance to these antibiotics in Gram-negative bacterial pathogens is the production of serine β-lactamase enzymes. A key aspect of the class A serine β-lactamase mechanism that remains unresolved and controversial is the identity of the residue acting as the catalytic base during the acylation reaction. Multiple mechanisms have been proposed for the formation of the acyl-enzyme intermediate that are predicated on understanding the protonation states and hydrogen-bonding interactions among the important residues involved in substrate binding and catalysis of these enzymes. For resolving a controversy of this nature surrounding the catalytic mechanism, neutron crystallography is a powerful complement to X-ray crystallography that can explicitly determine the location of deuterium atoms in proteins, thereby directly revealing the hydrogen-bonding interactions of important amino acid residues. Neutron crystallography was used to unambiguously reveal the ground-state active site protonation states and the resulting hydrogen-bonding network in two ligand-free Toho-1 β-lactamase mutants which provided remarkably clear pictures of the active site region prior to substrate binding and subsequent acylation [1,2] and an acylation transition-state analog, benzothiophene-2-boronic acid (BZB), which was also isotopically enriched with 11B. The neutron structure revealed the locations of all deuterium atoms in the active site region and clearly indicated that Glu166 is protonated in the BZB transition-state analog complex. As a result, the complete hydrogen-bonding pathway throughout the active site region could then deduced for this protein-ligand complex that mimics the acylation tetrahedral intermediate [3].

scholarly journals The Geometry of the Catalytic Active Site in [FeFe]-Hydrogenases is Determined by Hydrogen Bonding and Proton Transfer

2019 ◽  
Author(s):  
Jifu Duan ◽  
Stefan Mebs ◽  
Moritz Senger ◽  
Konstantin Laun ◽  
Florian Wittkamp ◽  
...  
Keyword(s):  
Hydrogen Bonding ◽  
Proton Transfer ◽  
Active Site ◽  
Time Resolved ◽  
X Ray Crystallography ◽  
Hydrogen Bonding Interactions ◽  
Catalytic Active Site ◽  
Catalytic Intermediates ◽  
Time Resolved Infrared Spectroscopy

The H2 conversion and CO inhibition reactivity of nine [FeFe]-hydrogenase constructs with semi-artificial cofactors was studied by in situ and time-resolved infrared spectroscopy, X-ray crystallography, and theoretical methods. Impaired hydrogen turnover and proton transfer as well as characteristic CO inhibition/ reactivation kinetics are assigned to varying degrees of hydrogen-bonding interactions at the active site. We show that the probability to adopt catalytic intermediates is modulated by intramolecular and protein-cofactor interactions that govern structural dynamics at the active site of [FeFe]-hydrogenases.<br>

Solid-State 17O NMR of Unstable Acyl-Enzyme Intermediates: A Direct Probe of Hydrogen Bonding Interactions in the Oxyanion Hole of Serine Proteases

2016 ◽  
Vol 120 (43) ◽  
pp. 11142-11150 ◽  
Author(s):  
Aaron W. Tang ◽  
Xianqi Kong ◽  
Victor Terskikh ◽  
Gang Wu
Keyword(s):  
Hydrogen Bonding ◽  
Solid State ◽  
Serine Proteases ◽  
17O Nmr ◽  
Oxyanion Hole ◽  
Hydrogen Bonding Interactions ◽  
Enzyme Intermediates

scholarly journals The molecular orbital study on the role of hydrogen bonding system in the active site of serine proteases.

1980 ◽  
Vol 28 (4) ◽  
pp. 1342-1344 ◽  
Author(s):  
SETSUKO NAKAGAWA ◽  
HIDEAKI UMEYAMA ◽  
TAKAKO KUDO
Keyword(s):  
Hydrogen Bonding ◽  
Molecular Orbital ◽  
Active Site ◽  
Serine Proteases ◽  
Bonding System ◽  
Molecular Orbital Study ◽  
Orbital Study ◽  
Hydrogen Bonding System

scholarly journals Joint neutron crystallographic and NMR solution studies of Tyr residue ionization and hydrogen bonding: Implications for enzyme-mediated proton transfer

2015 ◽  
Vol 112 (18) ◽  
pp. 5673-5678 ◽  
Author(s):  
Ryszard Michalczyk ◽  
Clifford J. Unkefer ◽  
John-Paul Bacik ◽  
Tobias E. Schrader ◽  
Andreas Ostermann ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Hydrogen Bonding ◽  
Carbonic Anhydrase ◽  
Proton Transfer ◽  
Active Site ◽  
Carbonic Anhydrase Ii ◽  
Hydrogen Bonding Interactions ◽  
Solution Studies ◽  
Hydrophilic Residues ◽  
Reversible Hydration

Human carbonic anhydrase II (HCA II) uses a Zn-bound OH−/H2O mechanism to catalyze the reversible hydration of CO2. This catalysis also involves a separate proton transfer step, mediated by an ordered solvent network coordinated by hydrophilic residues. One of these residues, Tyr7, was previously shown to be deprotonated in the neutron crystal structure at pH 10. This observation indicated that Tyr7 has a perturbed pKa compared with free tyrosine. To further probe the pKa of this residue, NMR spectroscopic measurements of [13C]Tyr-labeled holo HCA II (with active-site Zn present) were preformed to titrate all Tyr residues between pH 5.4–11.0. In addition, neutron studies of apo HCA II (with Zn removed from the active site) at pH 7.5 and holo HCA II at pH 6 were conducted. This detailed interrogation of tyrosines in HCA II by NMR and neutron crystallography revealed a significantly lowered pKa of Tyr7 and how pH and Tyr proximity to Zn affect hydrogen-bonding interactions.

Probing Hydrogen-Bonding Interactions in the Active Site of Medium-Chain Acyl-CoA Dehydrogenase Using Raman Spectroscopy†

Biochemistry ◽  
2003 ◽  
Vol 42 (40) ◽  
pp. 11846-11856 ◽  
Author(s):  
Jiaquan Wu ◽  
Alasdair F. Bell ◽  
Lian Luo ◽  
Avery W. Stephens ◽  
Marian T. Stankovich ◽  
...  
Keyword(s):  
Raman Spectroscopy ◽  
Hydrogen Bonding ◽  
Active Site ◽  
Medium Chain ◽  
Hydrogen Bonding Interactions ◽  
Chain Acyl
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