Enzymes
X-ray structure of perdeuterated DFPase – perdeuteration of enzymes for neutron diffraction
Perdeuteration of proteins is a much discussed strategy to overcome the problems of incoherent scattering caused by hydrogen in neutron diffraction experiments because deuterium displays a significantly lower incoherent scattering cross section compared to the normal hydrogen isotope. Expression of the protein in perdeuterated media is required for this. We report the X-ray structure of perdeuterated DFPase, which displays practically no differences to the hydrogenous structure. B-factors and RMSD values are reported. Even though a very big perdeuterated DFPase crystal was grown, it did not diffract neutrons. Reasons for this unexpected behaviour are discussed. The structure is presented in a new publication in Acta Cryst. F.
Characterization of the catalytic calcium binding-site in DFPase and comparison with other beta-propeller enzymes
The catalytic calcium binding-site of the enzymes DFPase displays remarkable similarities to metal binding sites in structurally related proteins like Paraoxonase (PON1), Drug Resistance Protein 35 (Drp35) from S. aureus or the Gluconolactonase XC5397 from Xanthomonas campestris. DFPase mutants targeting calcium binding residues and their structural characterization allow new insights with respect to metal binding and catalytic activity. The results are described in a new publication in Chemico-Biological Interactions (CBI). The article is part of a Special Issue of CBI on the occasion of the 10th International Meeting on Cholinesterases, which took place in Croatia in September 2009.
“Research Highlight” in Nature Chemistry
Our recently published article in JACS on engineering enantioselectivity in DFPase is now a “Research Highlight” in Nature Chemistry. The article is publicly available but a nature account is required.
Reversed Enantioselectivity of DFPase by Rational Design
Based on structural and mechanistic knowledge, mutants of the phosphotriesterase enzmye DFPase were created that reverse the enantioselectivity of the enzyme from the less toxic to the more toxic stereoisomer of G-type nerve agents. The mutants not only feature a reversed enantioselectivity but also increased enzymatic activity compared to the wildtype. The results are described in a new paper published in the Journal of the American Chemical Society (JACS).
Neutron Structure of the Enzyme DFPase
A new publication in the journal Proceedings of the National Academy of Sciences of the USA (PNAS) describes the neutron diffraction structure of the enzyme DFPase. Neutron diffraction allows the visualization of hydrogen atoms in protein structures and thus the determination of protonation states and orientations of solvent water molecules. The results of the study confirm the proposed reaction mechanism for DFPase and permit the directed introduction of modifications to the enzyme by mutagenesis to enhance both turnover rates and substrate diversity.
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