Lysozyme polypeptide chain

Lysozyme from bacteriophage T4 is a 164 amino acid polypeptide chain that folds into two domains (Figure 17.3) There are no disulfide bridges the two cysteine residues in the amino acid sequence, Cys 54 and Cys 97, are far apart in the folded structure. The stability of both the wild-type and mutant proteins is expressed as the melting temperature, Tm, which is the temperature at which 50% of the enzyme is inactivated during reversible beat denat-uration. For the wild-type T4 lysozyme the Tm is 41.9 °C. 

Figure 17.3 The polypeptide chain of lysozyme fiom hacteiiophage T4 folds into two domains. The N-terminal domain is of the a + P type, built up from two a helices (red) and a four-stranded antiparallel P sheet (green). The C-terminal domain comprises seven short a helices (brown and blue) in a rather irregular arrangement. (The last half of this domain is colored blue for clarity.)...

No sequence homologies can be detected. This is, perhaps, not surprising. The X-ray structure analysis of lysozyme by Phillips has shown that the polypeptide chain is folded in a way which puts none of the amino acids in sequential vicinity of the catalytic Asp-52 and Glu-37 that are near to the bound substrate. Comparable folding patterns can probably be realized with widely differing arrangements of amino acids, and thus the apparent lack of homologies. 

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... 

Fig. 2. —Atomic Arrangement in the Lysozyme Molecule in the Neighborhood of the Cleft with a Hexa-N-acetylchitohexaose Molecule Shown Bound to the Enzyme. (The main polypeptide chain is shown speckled, and NH and CO are indicated by line and full shading, respectively. Sugar residues A, B, and C are as observed in the binding of tri-N-acetylchitotriose. Residues D, E, and F occupy positions inferred from model building. It is suggested that the linkage hydrolyzed by the action of the enzyme is between residues D and E.) [Reprinted from Brookhaven Symp. Biol., 21, 120 (1968).]...

Fig. 2. Temperature dependence of the partial specific heat capacity for pancreatic ribonuclease A (RNase), hen egg-white lysozyme (Lys), sperm whale myoglobin (Mb), and catalase from Thermus thermophilus (CTT). The flattened curves are for RNase and Lys with disrupted disulfide cross-links and for apomyoglobin, when polypeptide chains have a random coil conformation without noticeable residual structure (Privalov et al., 1988).

Hen egg lysozyme is a small enzyme, having a single polypeptide chain of 129 aminoacid residues and a molecular mass of approximately 14400 Da, that catalyzes the hydrolysis of glycosidic linkages in bacterial cell walls. The schematic illustration of the active sites of lysozyme is shown in Fig. 10.29. 

Fig. 13. The conformation of polypeptide chain of hen egg white lysozyme as deduced from X-ray studies at 2 A resolution. The loop region is indicated by a circle. From Blake et...

The lysozyme molecule was determined to have three antigenic sites with residues coming from widely separated portions of the polypeptide chain the residues proposed as contacting and those synthesized to produce a linear sequence considered as best simulating the active site are seen in Fig. 15. In some instances the peptide synthesized in the reverse direction —for example, using the C-terminal amino acid of the hypothesized determinant as the amino terminus—was used as a control. In some instances, the sequence was considered to have directionality whereas in others it did not. 

Our understanding of the principles underlying the catalytic activity of enzymes has increased greatly in recent years. Enzymes are proteins and we know now that the polypeptide chain of a globular protein molecule will assume spontaneously a well-defined conformation (3). This native conformation of the macromolecule is essential to the function of the enzyme. Recent studies show that enzyme crystals, into which substrate molecules may penetrate by diffusion, have similar catalytic characteristics as enzyme solutions (4) and this result tends to strengthen our belief that the conformation of the enzyme molecule, as deduced from X-ray diffraction studies of enzyme crystals, is identical, or at least very similar, to the conformation responsible for the catalytic activity under physiological conditions. The structures of some enzymes, e.g., lysozyme (5), ribonuclease (6), and carboxypeptidase A (7), have been determined they are all consistent with the general belief that... 

Figure 3 Hen egg white lysozyme has 4 disulfide bridges. However, only one of them is easily reduced A marked with an arrow. The detail of its structure Is shown In BJ The disulfide radical anion is stabilized by interaction with the charged end of an Arginine residue (In green). In white the distances (In A) between the central carbon atom and the sulphur atoms. In red the polypeptidic chain.

Figure 8.7 Active sites may include distant residues. (A) Ribbon diagram of the enzyme lysozyme with several components of the active site shown in color. (B) A schematic representation of the primary structure of lysozyme shows that the active site is composed of residues that come from different parts of the polypeptide chain, [Drawn from 6LYZ.pdb.]...

The first protein crystal structure, myoglobin, was solved in 1960. The second, lysozyme, followed in 1965. In 1967 three structures were solved ribonuclease, chymotrypsin and carboxypeptidase. Thereafter, the number solved has increased almost exponentially year by year so that by 1979 there were some 161 structures known, at least at the level of tracing the fold of the polypeptide chain [6]. To date, there are well over 200 structures solved, but this number includes several structures of the same protein in a different crystal form or from a different species. Some protein structures are illustrated in Figure 1. 

A prerequisite for the catalytic function of an enzyme is its native tertiary structure which is determined by the number and sequence of amino acids (primary structure) forming the molecule. Favoured by hydrogen bonds, parts of the polypeptide chain exist in an a-helical or a (3-sheet structure (secondary structure). Most enzymes are globular proteins, the tertiary structure of which may be fixed by disulfide bonds between cysteine residues. A famous example is lysozyme (Fig. 20), consisting of 129 amino acids. A defined three-dimensional structure is... 

Materials. Proteins. The preparation of l-14C-acetyl derivatives of bovine /3-casein A and hen egg white lysozyme has been described (12, 13). Modification of /3-casein involved reaction of the protein with l-14C-acetic anhydride so that two lysine residues along the disordered, single polypeptide chain of 209 amino acids (11) were acetylated. A similar procedure was done with lysozyme so that one or two lysine residues on the exterior of the globular protein (14) were acetylated. These modifications ensured that the derivatives which were isolated in the lyophilized state had suitable specific activities (typically 1.5 juC/mg dry protein). The modification slightly increased the surface activity of lysozyme but not that of /3-casein (12, 13) in this paper the l-14C-acetyl derivatives are referred to as /3-casein and lysozyme. 

Figure 3 shows typical binding isotherms for hen egg lysozyme on binding sodium n-dodecylsulphate (SDS) in acid solution, pH 3.2, 25°C. Lysozyme (molecular mass 14,306) has a single polypeptide chain with an /V-terminal lysine residue, four disulphide bonds, and 18 cationic residues (11 arginyl, 6 lysyl, and 1 histidyl). At very low free SDS concentrations the binding isotherms are very steep but... 

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