Current status of neutron crystallography in structural biology

Abstract

Hydrogen atoms and hydration water molecules in proteins are essential for many biochemical processes, especially enzyme catalysis. Neutron crystallography enables direct observation of hydrogen atoms, and reveals molecular recognition through hydrogen bonding and catalytic reactions involving proton-coupled electron transfer. The use of neutron crystallography is still limited for proteins, but its popularity is increasing owing to an increase in the number of diffractometers for structural biology at neutron facilities and advances in sample preparation. According to the characteristics of the neutrons, monochromatic or quasi-Laue methods and the time-of-flight method are used in nuclear reactors and pulsed spallation sources, respectively, to collect diffraction data. Growing large crystals is an inevitable problem in neutron crystallography for structural biology, but sample deuteration, especially protein perdeuteration, is effective in reducing background levels, which shortens data collection time and decreases the crystal size required. This review also introduces our recent neutron structure analyses of copper amine oxidase and coppercontaining nitrite reductase. The neutron structure of copper amine oxidase gives detailed information on the protonation state of dissociable groups, such as the quinone cofactor, which are critical for catalytic reactions. Electron transfer via a hydrogen-bond jump and a hydroxide ion ligation in copper-containing nitrite reductase are clarified, and these observations are consistent with the results from the quantum chemical calculations. This review article is an extended version of the Japanese article, Elucidation of

Resveratrol Derivatives Inhibit Transthyretin Fibrillization: Structural Insights into the Interactions between Resveratrol Derivatives and Transthyretin

“…Therefore, the exchangeable hydrogen atoms bound to nitrogen and oxygen atoms could be observed as a mixture of hydrogen and deuterium atoms. Hydrogens and deuteriums can be distinguished due to their negative and positive neutron scattering lengths …”

“…Hydrogens and deuteriums can be distinguished due to their negative and positive neutron scattering lengths. 34 Clear positive peaks were observed at the oxygen atoms of the resorcinol group, indicating that these hydroxy groups donate hydrogen atoms to the hydrogen bond with S117 (Figure 8). S117 donated hydrogen atoms in the hydrogen bonding with the central water molecule, creating a network of hydrogen bonds.…”

Overlooked Hydrogen Bond in a Blue Copper Protein Uncovered by Neutron and Sub-Ångström Resolution X-ray Crystallography

“…Moreover, because the energy of neutrons with wavelengths used in typical macromolecular X-ray crystallography is very low (e.g., 81 meV [neutron] compared to 12 keV [X-ray] at λ = 1.0 Å), it is possible to determine metalloprotein structures without photoreduction with neutron crystallography. 19,20 Although a sub-ångstrom X-ray structure 21 and a neutron structure 22 of a BCP amicyanin from Paracoccus denitrificans (PdAmi) have been reported previously, the environments of the T1Cu sites differ from BCP to BCP and thus a comparative analysis of BCPs is necessary to fully understand the structure−function relationships of BCPs. In particular, differences in the ligand-containing loop (C−H-M loop) contribute to the chemical properties of the copper center.…”

“…H and its isotope deuterium (D) atoms contribute to neutron scattering to the same extent as C, N, and O atoms. Moreover, because the energy of neutrons with wavelengths used in typical macromolecular X-ray crystallography is very low (e.g., 81 meV [neutron] compared to 12 keV [X-ray] at λ = 1.0 Å), it is possible to determine metalloprotein structures without photoreduction with neutron crystallography. , …”

Description of peptide bond planarity from high-resolution neutron crystallography

“…However, due to their possession of only one electron, the determination of the coordinates using X-ray crystallography is difficult. Neutron crystallography leverages the interactions between neutron beams and atomic nuclei to determine the coordinates of hydrogen atoms with almost the same precision as those of other atoms such as carbon, nitrogen, and oxygen [ 1 , 2 ]. Moreover, in neutron crystallography, the hydrogen (protium)-deuterium (H/D) exchange ratio can provide valuable information for understanding the conformational dynamics and exposure to water [ 3 – 6 ].…”