Protein-protein interaction on lysozyme crystallization revealed by rotational diffusion analysis.
Intermolecular interactions between protein molecules diffusing in various environments underlie many biological processes as well as control protein crystallization, which is a crucial step in x-ray protein structure determinations. Protein interactions were investigated through protein rotational diffusion analysis. First, it was confirmed that tetragonal lysozyme crystals containing fluorescein-tagged lysozyme were successfully formed with the same morphology as that of native protein. Using this nondisruptive fluorescent tracer system, we characterized the effects of sodium chloride and ammonium sulfate concentrations on lysozyme-lysozyme interactions by steady-state and time-resolved fluorescence anisotropy measurements and the introduction of a novel interaction parameter, k(rot). The results suggested that the specific attractive interaction, which was reflected in the retardation of the protein rotational diffusion, was induced depending on the salt type and its concentration. The change in the attractive interactions also correlated with the crystallization/precipitation behavior of lysozyme. Moreover, we discuss the validity of our rotational diffusion analysis through comparison with the osmotic second virial coefficient, B(22), previously reported for lysozyme and those estimated from k(rot).
Takahashi
Daisuke
D
Institute of Biophysics, Faculty of Agriculture, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan.
Nishimoto
Etsuko
E
Murase
Tadashi
T
Yamashita
Shoji
S
D001665:Binding Sites; D003198:Computer Simulation; D003460:Crystallization / Q000379:methods*; D004058:Diffusion; D008856:Microscopy, Fluorescence / Q000379:methods*; D008956:Models, Chemical; D008958:Models, Molecular; D046912:Multiprotein Complexes / Q000737:chemistry / Q000648:ultrastructure; D009113:Muramidase / Q000737:chemistry* / Q000648:ultrastructure*; D011485:Protein Binding; D011487:Protein Conformation; D025941:Protein Interaction Mapping / Q000379:methods*
D046912:Multiprotein Complexes; D009113:Muramidase