Neutron crystallographic study on rubredoxin from Pyrococcus furiosus by BIX-3, a single-crystal diffractometer for biomacromolecules

The structure of a partially deuterated rubredoxin from the hyperthermophilic archaeon Pyrococcus furiosus, an organism that grows optimally at 100[degrees]C, was determined by using the neutron single-crystal diffractometer dedicated for biological macromolecules (BIX-3) at the JRR-3M reactor of the Japan Atomic Energy Research Institute. Data were collected at room temperature up to a resolution of 1.5 [Angstrom], and the completeness factor of the data set was 81.9%. The model contains 306 H and 50 D atoms. A total of 37 hydration water molecules were identified, with 15 having all three atoms fully located and the remaining [D.sub.2]O molecules partially defined. The model has been refined to final agreement factors of R = 18.6% and [R.sub.free] = 21.7%. Several orientations of the O-D bonds of side chains, whose assignments from x-ray data were previously ambiguous, were clearly visible in the neutron structure. Although most backbone N-H bonds had undergone some degree of H/D exchange throughout the rubredoxin molecule, 5 H atom positions still had distinctly negative (H) peaks. The neutron Fourier maps clearly showed the details of an extensive set of H bonds involving the [ND.sub.3.sup.+] terminus that may contribute to the unusual thermostability of this molecule.