title: Unambiguous determination of H-atom positions: comparing results from neutron and high-resolution X-ray crystallography.
abstract: The locations of H atoms in biological structures can be difficult to determine using X-ray diffraction methods. Neutron diffraction offers a relatively greater scattering magnitude from H and D atoms. Here, 1.65 A resolution neutron diffraction studies of fully perdeuterated and selectively CH(3)-protonated perdeuterated crystals of Pyrococcus furiosus rubredoxin (D-rubredoxin and HD-rubredoxin, respectively) at room temperature (RT) are described, as well as 1.1 A resolution X-ray diffraction studies of the same protein at both RT and 100 K. The two techniques are quantitatively compared in terms of their power to directly provide atomic positions for D atoms and analyze the role played by atomic thermal motion by computing the sigma level at the D-atom coordinate in simulated-annealing composite D-OMIT maps. It is shown that 1.65 A resolution RT neutron data for perdeuterated rubredoxin are approximately 8 times more likely overall to provide high-confidence positions for D atoms than 1.1 A resolution X-ray data at 100 K or RT. At or above the 1.0sigma level, the joint X-ray/neutron (XN) structures define 342/378 (90%) and 291/365 (80%) of the D-atom positions for D-rubredoxin and HD-rubredoxin, respectively. The X-ray-only 1.1 A resolution 100 K structures determine only 19/388 (5%) and 8/388 (2%) of the D-atom positions above the 1.0sigma level for D-rubredoxin and HD-rubredoxin, respectively. Furthermore, the improved model obtained from joint XN refinement yielded improved electron-density maps, permitting the location of more D atoms than electron-density maps from models refined against X-ray data only.
authors.affiliation: Oak Ridge National Laboratory, USA. anna.s.gardberg@gmail.com
authors.forename: Anna S
authors.initials: AS
authors.lastname: Gardberg
authors.forename: Alexis Rae
authors.initials: AR
authors.lastname: Del Castillo
authors.forename: Kevin L
authors.initials: KL
authors.lastname: Weiss
authors.forename: Flora
authors.initials: F
authors.lastname: Meilleur
authors.forename: Matthew P
authors.initials: MP
authors.lastname: Blakeley
authors.forename: Dean A A
authors.initials: DA
authors.lastname: Myles
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry*; D018360:Crystallography, X-Ray / Q000379:methods*; D033363:Neutron Diffraction / Q000379:methods; D020027:Pyrococcus furiosus / Q000737:chemistry*; D012416:Rubredoxins / Q000737:chemistry*; D014961:X-Ray Diffraction / Q000379:methods
chemical_list: D001426:Bacterial Proteins; D012416:Rubredoxins