Lysozyme domain motion

Hayward, S., Kitao, A., Berendsen, H.J.C. Model-free methods to analyze domain motions in proteins from simulation A comparison of normal mode analysis and molecular dynamics simulation of lysozyme. Proteins 27 (1997) 425-437. 

The internal motion of T4 lysozyme in the crystal was interpreted as an inter-domain motion corresponding to opening and closing of the active site cleft (Weaver et al., 1989). Hinge-bending and substrate-induced conformational transition in T4 lysozyme in solution were confirmed in a study by site-directed labelling (Mchaourban et al., 1997). 

Neutron spectroscopy is becoming a principal tool for the study of protein dynamics (Cusack, 1986, 1989 Middendorf, 1984 Middendorf et al., 1984). Current instruments cover motions with characteristic times from 10 to 10 sec. This range embraces essentially all protein modes excited at room temperature (the soft modes), including motions of the solvent shell and also low-frequency large-scale domain motions, like the hinge-bending motion of the lysozyme domains that form the... 

Lysozyme is a key system in the development of our understanding of the structure and function of proteins. It was the first enzyme whose x-ray structure was determined at high resolution (Blake et al. 1965), one of the earliest enzymes for which a detailed reaction mechanism was proposed and one of the test systems for molecular dynamics simulations. In this section, we review some of the simulation studies of lysozyme with emphasis on the reaction mechanism and on the large scale domain motion involved in the mechanism. 

Most of the information on interdomain motions come from high-resolution crystal structures several reviews are available (Janin and Wodak 1983 Bennett andHuber 1984 Gerstein et al. 1994). Calculations ofhinge bending modes and domain motions in proteins other than lysozyme have been made. They include antibody molecules where the interdomain motions occur on a nanosecond time scale (McCammon and Karplus 1977 Oi et al. 1984), 1-arabinose-binding protein (Mao et al. 1982), liver alcohol dehydrogenase (Colona-Cesari et al. 1986) and the mouse... 

In an early study of lysozyme ([McCammon et al. 1976]), the two domains of this protein were assumed to be rigid, and the hinge-bending motion in the presence of solvent was described by the Langevin equation for a damped harmonic oscillator. The angular displacement 0 from the equilibrium position is thus governed by... 

One of the main attractions of normal mode analysis is that the results are easily visualized. One can sort the modes in tenns of their contributions to the total MSF and concentrate on only those with the largest contributions. Each individual mode can be visualized as a collective motion that is certainly easier to interpret than the welter of information generated by a molecular dynamics trajectory. Figure 4 shows the first two normal modes of human lysozyme analyzed for their dynamic domains and hinge axes, showing how clean the results can sometimes be. However, recent analytical tools for molecular dynamics trajectories, such as the principal component analysis or essential dynamics method [25,62-64], promise also to provide equally clean, and perhaps more realistic, visualizations. That said, molecular dynamics is also limited in that many of the functional motions in biological molecules occur in time scales well beyond what is currently possible to simulate. 

Figure 4 DynDom [67] analysis of the first two normal modes of human lysozyme. Dark grey and white indicate the two dynamic domains, separated by the black hinge bending region. The vertical line represents a hinge axis that produces a closure motion in the first normal mode. The horizontal line represents a hinge axis that produces a twisting motion in the second normal mode. (Adapted from Ref. 68.) The DynDom program is available from the Internet at http //md. chem.rug.nl/ steve/dyndom.html.

Harvey and Hoekstra (1972) determined the dielectric constant and loss for lysozyme powders as a function of hydration level in the frequency range 10 —10 Hz. At water contents less than 0.3 h, they found a dispersion at 170 MHz, which increased somewhat with increasing hydration, and a new dispersion at about 10 Hz that develops at high hydration. These dispersions, detected by time-domain techniques, remain measurable down to the lowest temperature studied, — 60°C. Water mobility in the hydration shell below 0 C is in line with other observations of nonfreezing water. Above 0.3 h, in the stage of the hydration process at which condensation completes the surface monolayer, water motion increased strongly with increased hydration (Fig. 11). 

E-fS ES ES. The standard deviation of the distribution, (Atopen ) = 8.3 2ms, reflects the distribution bandwidth. For the individual T4 lysozyme molecules examined under the same enz unatic reaction conditions, we found that the first and second moments of the single-molecule topen distributions are homogeneous, within the error bars. The hinge-bending motion allows sufficient structural flexibility for the enzyme to optimize its domain conformation the donor fluorescence essentially reaches the same intensity in each turnover, reflecting the domain conformation reoccurrence. The distribution with a defined first moment and second moment shows typical oscillatory conformational motions. The nonequilibrium conformational motions in forming the active enzymatic reaction intermediate states intrinsically define a recurrence of the essentially similar potential surface for the enzymatic reaction to occur, which represents a memory effect in the enzymatic reaction conformational dynamics [12,41,42]. 

All proteins are flexible. We can see flexibility even in cytochrome C on changing the iron atom from Fe to Fe. The flexibility extends to the surface. In lysozyme the protein closes on the substrate and there are small motions throughout the two domains. The question is therefore about the value of such movements, since all proteins move in all reactions. In other words we must try to define the functional value of mobihty, i.e. its purpose . 

Mouse epidermal growth factor (EOF) is a relatively small protein consisting of 53 amino acids with three disulfide bonds. EGF can be described as a right-handed mitten where the thumb and palm represent separate domains and the hollow is postulated as the site where the EGF receptor binds. Ikura and Go have carried out a normal mode analysis of this protein. As was the case with lysozyme, the lowest frequency mode (4.1 cm" ) corresponds to a hinge-bending motion. This finding helps to rationalize the difference between two NMR structures which have been solved for this molecule. These two structures differed in terms of the distance between the two domains, due to the paucity of interdomain NOEs. Normal modes have also been incorporated into the structure determination process, as described in Sections 3.4 and 3.5. 

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