A Neutron crystallographic analysis of a rubredoxin mutant at 1.6 resolution

Chatake, Toshiyuki*; Kurihara, Kazuo; Tanaka, Ichiro*; Tsyba, I.*; Bau, R.*; Jenney, F. E. Jr.*; Adams, M. W. W.*; Niimura, Nobuo

A neutron diffraction study has been carried out at 1.6 resolution on a mutant rubredoxin from using the BIX-3 single-crystal diffractometer at the JRR-3 reactor of JAERI. In order to study the unusual thermostability of rubredoxin from , the hydrogen-bonding patterns were compared between the native and a 'triple-mutant' variant where three residues were changed so that they are identical to those in a mesophilic rubredoxin. In the present study, some minor changes were found between the wild-type and mutant proteins in the hydrogen-bonding patterns of the Trp3/Tyr3 region. The H/D-exchange ratios in the protein were also studied. The results suggest that the backbone amide bonds near the four Cys residues of the FeS redox center are most resistant to H/D exchange. In addition, the 1.6 resolution of the present neutron structure determination has revealed a more detailed picture than previously available of some portions of the water structure, including ordered and disordered O-D bonds.