On the conformation of the hen egg-white lysozyme molecule

Crystals of hen egg-white lysozyme, grown at pH 4.7 (Alderton & Fevold 1946), are tetragonal with a = b = 79.1 Å, c = 37.9 Å, space group P 4 3 2 1 2 (Palmer, Ballantyre & Galvin 1948; Blake, Fenn, North, Phillips & Poljak 1962). Each of the eight asymmetric units in the cell comprises a single lysozyme molecule, molecular weight about 14 600, together with 1 M sodium chloride solution which constitutes some 33.5% of the weight of the crystal (Steinrauf 1959). The structure of these crystals has been determined by X-ray analysis by the method of multiple isomorphous replacement developed in the studies of haemoglobin (Green, Ingram & Perutz 1954; Blow 1958; Perutz, Rossmann, Cullis, Muirhead, Will & North 1960) and myoglobin (Kendrew, Dickerson, Strandberg, Hart, Davies, Phillips & Shore 1960). Anomalous scattering data were used in conjunction with the isomorphous replacement intensity differences (North 1965) to form a joint probability distribution for the phase of each reflexion. The position of the centroid of each probability distribution gave a phase angle and weighting factor for each reflexion from which the electron density map with minimum r.m.s. error was calculated (Blow & Crick 1959). A large number of different heavy atom derivatives were studied (Poljak 1963; Blake, Koenig, Mair, North, Phillips & Sarma 1965) and three proved satisfactory for calculating an electron density map at 2 Å resolution. They contained respectively ortho -mercuri hydroxytoluene para -sulphonic acid, UO 2 F 5 3- and an ion derived from UO 2 (NO 3 ) 2 , probably UO 2 (OH) n (n-2)-

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The active site of hen egg-white lysozyme: flexibility and chemical bonding

Acta Crystallographica Section D Biological Crystallography ◽

10.1107/s1399004714001928 ◽

2014 ◽

Vol 70 (4) ◽

pp. 1136-1146 ◽

Cited By ~ 17

Author(s):

Jeanette Held ◽

Sander van Smaalen

Keyword(s):

Quantum Theory ◽

Electron Density ◽

Active Site ◽

Egg White ◽

Hen Egg White Lysozyme ◽

Acta Cryst ◽

Covalent Bonds ◽

Egg White Lysozyme ◽

Hen Egg White ◽

Hen Egg

Chemical bonding at the active site of hen egg-white lysozyme (HEWL) is analyzed on the basis of Bader's quantum theory of atoms in molecules [QTAIM; Bader (1994),Atoms in Molecules: A Quantum Theory.Oxford University Press] applied to electron-density maps derived from a multipole model. The observation is made that the atomic displacement parameters (ADPs) of HEWL at a temperature of 100 K are larger than ADPs in crystals of small biological molecules at 298 K. This feature shows that the ADPs in the cold crystals of HEWL reflect frozen-in disorder rather than thermal vibrations of the atoms. Directly generalizing the results of multipole studies on small-molecule crystals, the important consequence for electron-density analysis of protein crystals is that multipole parameters cannot be independently varied in a meaningful way in structure refinements. Instead, a multipole model for HEWL has been developed by refinement of atomic coordinates and ADPs against the X-ray diffraction data of Wang and coworkers [Wanget al.(2007),Acta Cryst.D63, 1254–1268], while multipole parameters were fixed to the values for transferable multipole parameters from the ELMAM2 database [Domagalaet al.(2012),Acta Cryst.A68, 337–351] . Static and dynamic electron densities based on this multipole model are presented. Analysis of their topological properties according to the QTAIM shows that the covalent bonds possess similar properties to the covalent bonds of small molecules. Hydrogen bonds of intermediate strength are identified for the Glu35 and Asp52 residues, which are considered to be essential parts of the active site of HEWL. Furthermore, a series of weak C—H...O hydrogen bonds are identified by means of the existence of bond critical points (BCPs) in the multipole electron density. It is proposed that these weak interactions might be important for defining the tertiary structure and activity of HEWL. The deprotonated state of Glu35 prevents a distinction between the Phillips and Koshland mechanisms.

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An X-ray study of the physiological-temperature form of hen egg-white lysozyme at 2 Å resolution

Proceedings of the Royal Society of London Series B Biological Sciences ◽

10.1098/rspb.1983.0021 ◽

1983 ◽

Vol 217 (1209) ◽

pp. 471-489 ◽

Cited By ~ 29

Keyword(s):

High Temperature ◽

Egg White ◽

Hen Egg White Lysozyme ◽

Final Model ◽

Egg White Lysozyme ◽

Isomorphous Replacement ◽

Tetragonal Form ◽

Hen Egg White ◽

Mean Square Difference ◽

Hen Egg

The structure of the high-temperature orthorhombic form of hen egg-white lysozyme has been determined at 2.0 Å resolution. Initial images of the molecule were obtained at 6.0 Å resolution both by double isomorphous replacement and by molecular replacement with use of the known structure of the room-temperature tetragonal lysozyme. The initial model thus obtained ( R = 0.52 at 6.0 Å) was refined first as a rigid body at 6.0 Å and then by restrained least squares at 2.5 Å and later at 2.0 Å resolution. The final model ( R = 0.23 at 2.0 Å) was compared with that of the tetragonal form: the structures are very similar with a root mean square difference in superimposed α-carbon coordinates of 0.46 Å. There are, however, differences which are caused by a crystal contact involving the upper part of this active site in the high-temperature orthorhombic form. Because of this, residues Trp 62 and Pro 70 are much better ordered than in the tetragonal form, where they are exposed to solvent. These differences can partly explain the difficulty of inhibitor-binding in high-temperature orthorhombic crystals, but do not seem to reflect the particular behaviour of lysozyme in solution at high temperature.

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