Lysozyme hydration

There have been two different interpretations of the slow dynamics observed in the SD of the lysozyme hydration layer. The first attributes the intermediate time-scales (30 0 ps) to slow water. Bagchi and co-workers employed the dynamic exchange model to relate the observed slow dynamics to the timescale of the fluctuation of water in the hydration layer [11]. In an alternative interpretation. Song et al. used the formulation developed by Song and Marcus that relates the solvation time correlation function to the DR of the medium. They attributed the... 

Figure 25. Arrhenius plot of the /D for lysozyme hydration water, calculated from MD simulations. The curve shows an high-T dy namic crossover similar to the one observed hy QENS (Fig. 24) [74].

Recently Blake et al.153) made such studies in the case of human (HL) and tortoise egg-white (TEWL) lysozyme based on crystallographic refinements at 1,5 and 1,6 A resolution, respectively. By these investigations they attempted to obtain information on the perturbations of water structure in the hydration shell by neighboured protein molecules and by high salt concentrations as well as on the degree of order of the bound water. The authors came to the conclusion that the number of ordered water molecules are 128 in TEWL and 140 in HL, whereas the overall content is made up of 650 and 350 water molecules per lysozyme molecule. 

Carbon atom, 4. See also Atomic orbitals Carbon dioxide hydration, 197-199. See also Carbonic anhydrase Carbonic anhydrase, 197-199,200 Carbonium ion transition state, 154, 159 Carboxypeptidase A, 204-205 Catalysis, general acid, 153,164,169 in carboxypeptidase A, 204-205 free energy surfaces for, 160, 161 in lysozyme, 154... 

The ROA spectra of native and prehbrillar amyloidogenic human lysozyme are displayed in Figure 7, together with a MOLSCRIPT diagram of the native structure. The ROA spectrum of the native protein is very similar to that of hen lysozyme (Fig. 5). However, large changes have occurred in the ROA spectrum of the prehbrillar intermediate. In particular, the positive 1340 cm-1 ROA band assigned to hydrated... 

It has been suggested recently that PPII helix may be the killer conformation in such diseases (Blanch et al., 2000). This was prompted by the observation, described in Section III,B, of a positive band at 1318 cm-1, not present in the ROA spectrum of the native state, that dominates the ROA spectrum of a destabilized intermediate of human lysozyme (produced by heating to 57°C at pH 2.0) that forms prior to amyloid fibril formation. Elimination of water molecules between extended polypeptide chains with fully hydrated 0=0 and N—H groups to form... 

Remmele et al. [3.55] studied with infrared spectroscopy the structure-hydration behavior of a 49.4 mg/mL lysozyme D20 solution with and without 10 % sucrose. The sample was cooled n in the measuring chamber to -100 °C and then connected to a freeze drying installation, after which the temperature of the sample was raised to +40 °C ... 

Remmele, R. L., Stushoff, Carpenter, J. F. Real-time spectroscopy analysis of lysozyme during Lyophilization structure-hydration behavior and influence of sucrose. American Chemical Society Symposium, Ser. 567 (Formulation and delivery of proteins and peptides) 1994. 1994 American Chemical Society... 

Fig. 18. The proton spin-lattice relaxation rate recorded as a function of the magnetic field strength plotted as the proton Larmor frequency for lysozyme samples. Dry ( ), hydrated to 8.9% ( ), 15.7% (O). 23.1% (A), and cross-linked in a gel ( ). The solid lines were computed from the theory. The solid lines are fits to the data using Eq. (4) with Rs given by Eq. (6). The two parameters adjusted are Rsl and b (97). The small peaks most apparent in the dry samples are caused by cross-relaxation to the peptide nitrogen spin (90,122).

Disposable cuvettes are used only once to eliminate the possibility of lysozyme carryover. All microcentrifuge tubes and pipette tips are autoclaved, and buffers and BSA solutions are not used beyond 7 days. The M. lysodeikticus suspension is prepared 18-24 hr before use and set at 37° with shaking, to hydrate fully. It was found that substrate prepared just before use settled much faster and interfered with the signal at the low concentrations of enzyme used. 

It is worthwhile to present some examples of the types of results available from Monte Carlo simulations of peptide solvent systems. In Figure 6 we present the convergence of the energy of hydration of the lysozyme crystal over a million configurations. 

Figure 6. The average energy of the water of hydration of the triclinic lysozyme crystal as a function of the number of configurations generated in the Monte Carlo simulation. The upper curve (A) corresponds to the cumulative statistical average, and the lower curve fU gives the statistical average over sequential sets of 5,000...

Fig. 10.3. Water molecules in a cavity of lysozyme. Only the surrounding residues are displayed. The isosurfaces of water oxygen (green) and hydrogen (pink) for the 3D distributions larger than 8 (left), the most probable model of the hydration structure reconstructed from the isosurface plots (center), and the crystallographic...

This corresponds to a hydration level of 0.31 g water g-1 protein, a value very close to that obtained experimentally for lysozyme (see Ihble 23.1). If we add another 130-150 water molecules to cover the lysozyme surface completely, a value of 0.46 g water g-1 protein is reached. In this range, IR and NMR experiments indicate a discontinuity suggestive of another kind of more mobile and less tightly bound water. 

The hydration of a protein can be described in several steps [622, 810, 827-829]. If the molar ratio water/lysozyme is gradually increased, a number of discrete levels of hydration are observed [810, 828], as established by a variety of methods summarized in Fig. 23.1 and in Ihble 23.2 ... 

Table 23.2. Steps in the hydration of lysozyme. The hydration level is given in g water g-1 protein, and numbers in parentheses refer to number of water ... 

Hydration of an alanine side-chain. In human lysozyme, the methyl group of Ala92 is surrounded by four water molecules located on a semi-circle ([625], Fig. 23.6). These hydration waters are buried in the protein which might be the reason why they are so well ordered. We can also assume that such hydration schemes occur at the protein periphery but they are not seen in the X-ray analyses due to larger-thermal motion and/or disorder of the water molecules. 

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