Publications (45) View all
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Article: Preliminary time-of-flight neutron diffraction study of human deoxyhemoglobin.
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ABSTRACT: Human hemoglobin (HbA) is an intricate system that has evolved to efficiently transport oxygen molecules (O(2)) from lung to tissue. Its quaternary structure can fluctuate between two conformations, T (tense or deoxy) and R (relaxed or oxy), which have low and high affinity for O(2), respectively. The binding of O(2) to the heme sites of HbA is regulated by protons and by inorganic anions. In order to investigate the role of the protonation states of protein residues in O(2) binding, large crystals of deoxy HbA (approximately 20 mm(3)) were grown in D(2)O under anaerobic conditions for neutron diffraction studies. A time-of-flight neutron data set was collected to 1.8 A resolution on the Protein Crystallography Station (PCS) at the spallation source run by Los Alamos Neutron Science Center (LANSCE). The HbA tetramer (64.6 kDa; 574 residues excluding the four heme groups) occupies the largest asymmetric unit (space group P2(1)) from which a high-resolution neutron data set has been collected to date.Acta Crystallographica Section F Structural Biology and Crystallization Communications 05/2008; 64(Pt 4):270-3. · 0.51 Impact Factor -
Article: Recent results on hydrogen and hydration in biology studied by neutron macromolecular crystallography.
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ABSTRACT: Neutron diffraction provides an experimental method of directly locating hydrogen atoms in proteins, a technique complimentary to ultra-high-resolution [1, 2] X-ray diffraction. Three different types of neutron diffractometers for biological macromolecules have been constructed in Japan, France and the United States, and they have been used to determine the crystal structures of proteins up to resolution limits of 1.5-2.5 A. Results relating to hydrogen positions and hydration patterns in proteins have been obtained from these studies. Examples include the geometrical details of hydrogen bonds, H/D exchange in proteins and oligonucleotides, the role of hydrogen atoms in enzymatic activity and thermostability, and the dynamical behavior of hydration structures, all of which have been extracted from these structural results and reviewed. Other techniques, such as the growth of large single crystals, the preparation of fully deuterated proteins, the use of cryogenic techniques, and a data base of hydrogen and hydration in proteins, will be described.Cellular and Molecular Life Sciences CMLS 03/2006; 63(3):285-300. · 6.57 Impact Factor -
Article: Crystallization of a large single crystal of cubic insulin for neutron protein crystallography.
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ABSTRACT: The growth of a large single crystal of cubic porcine insulin for characterization of hydrogen and hydration in cubic insulin crystals by neutron diffraction analysis is reported. Growth in D2O was investigated based on the phase diagram for cubic insulin to determine appropriate growth conditions, and a large single crystal was then successfully grown by a dialysis method to a size of 4.0 x 4.0 x 1.3 mm3. Neutron diffraction analysis of the cubic insulin crystals was carried out using a single-crystal diffractometer at the JRR-3M reactor of the Japan Atomic Energy Research Institute. In preliminary analysis, Npi appears to be protonated and Ntau deprotonated in His5 in the B-chain, whereas both Npi and Ntau are protonated in His10.Journal of Synchrotron Radiation 02/2004; 11(Pt 1):41-4. · 2.73 Impact Factor -
Article: A neutron crystallographic analysis of a rubredoxin mutant
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ABSTRACT: In order to study the unusual thermostability of rubredoxin from Pyrococcus furiosus (RubPf), the structure of a 'triple mutant' of this rubredoxin, which is less thermostable than the wild type, was solved at 1.5- resolution by neutron-diffraction analysis using the BIX-3 diffractometer at the JRR-3M reactor of JAERI. The positions of the non-hydrogen atoms are almost the same as the native rubredoxin; however, for Trp3 M Tyr3, large structural changes were found, so that their hydrogen-bonding schemes are significantly different. Some positions of the hydrogen atoms and molecules of hydration are shifted in certain regions, suggesting that such differences may contribute to the differences in thermostability between this 'triple-mutant' rubredoxin and wild-type rubredoxin.Applied Physics A 01/2002; 74:s1280-s1282. · 1.63 Impact Factor -
Article: High-resolution X-ray study of the effects of deuteration on crystal growth and the crystal structure of proteinase K.
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ABSTRACT: Deuteration of macromolecules is an important technique in neutron protein crystallography. Solvent deuteration of protein crystals is carried out by replacing water (H(2)O) with heavy water (D(2)O) prior to neutron diffraction experiments in order to diminish background noise. The effects of solvent deuteration on the crystallization of proteinase K (PK) with polyethylene glycol as a precipitant were investigated using high-resolution X-ray crystallography. In previous studies, eight NO(3)(-) anions were included in the PK crystal unit cell grown in NaNO(3) solution. In this study, however, the PK crystal structure did not contain NO(3)(-) anions; consequently, distortions of amino acids arising from the presence of NO(3)(-) anions were avoided in the present crystal structures. High-resolution (1.1 Å) X-ray diffraction studies showed that the degradation of PK crystals induced by solvent deuteration was so small that this degradation would be negligible for the purpose of neutron protein crystallography experiments at medium resolution. Comparison of the nonhydrogen structures of nondeuterated and deuterated crystal structures demonstrated very small structural differences. Moreover, a positive correlation between the root-mean-squared differences and B factors indicated that no systematic difference existed.Acta Crystallographica Section F Structural Biology and Crystallization Communications 11/2011; 67(Pt 11):1334-8. · 0.51 Impact Factor
