X-RAY DIFFRACTION STUDIES OF HYDROGEN BONDING PATTERNS IN PROTEIN STRUCTURES AT ATOMIC RESOLUTION

Alexander N. Popov¹, Galina S. Kachalova² & Hans D. Bartunik²

¹EMBL, c/o DESY, Notkestraße 85, 22603 Hamburg, Germany;
² Max-Planck-AG Proteindynamik, MPG-ASMB, c/o DESY, Notkestraße 85, 22603 Hamburg, Germany.
e-mail: sasha@embl-hamburg.de;

Keywords : protein, X-ray crystal structure, atomic resolution, hydrogen bond

We analysed hydrogen bonding patterns in protein X-ray crystal structures at atomic resolution around 1.0 A. The models were refined using SHELXL-97 [1] employing restrained anisotropic atomic displacement parameters and calculated hydrogen positions according to geometrical criteria. In a first step of our study we have intended to examine the grounds of hydrogen atoms inclusion in protein model structure at atomic resolution. Due to the high quality and contrast of electron density maps a lot of hydrogen atoms can be located directly. A statistical analysis has revealed the coincidence of experimentally observed peaks in omit or difference density maps with the calculated hydrogen atoms positions. We derived that the probability of visualibility of hydrogen atoms depends on the displacement parameters of the 'heavy atoms' to which they are bound and on the noise level of difference maps. The potential of using X-ray diffraction for locating hydrogens in protein structures is compared to other techniques e.g. neutron diffraction.

Analysis of hydrogen bond patterns and their metrics of the strong and weak bonding interactions was made for these protein structures. All potential donor and acceptor peptide groups turned out to be mainly engaged in strong hydrogen bonding interactions. Taking into account of weak hydrogen bonding interactions nearly all C=O peptide groups, even those in b-sheets realized the hydrogen bonding potential of lone pairs on the carbonyl oxygen. A directionality of these hydrogen bonding differs between protein and small molecule crystal structures [2]. When comparing protein structures in different crystal forms, we observed that the conformation of the hydrogen bonding patterns was substantially more conserved than the conformation of the protein backbone . We propose this conservation of the overall hydrogen bonding pattern to have functional significance.

[1] Sheldrick G.W., Gottingen University SHELX-97 (1997)
[2] Taylor R., Kennard O., Versichel W., J.Am.Chem.Soc. 105 (1983) 5761-5766