Lysozyme enzyme efficiency

The term represents the kinetic efficiency of the enzyme. Table 14.4 lists turnover numbers for some representative enzymes. Catalase has the highest turnover number known each molecule of this enzyme can degrade 40 million molecules of HgOg in one second At the other end of the scale, lysozyme requires 2 seconds to cleave a glycosidic bond in its glycan substrate. 

Disruption of microbial cells is rendered difficult due to the presence of the microbial cell wall. Despite this, a number of very efficient systems exist that are capable of disrupting large quantities of microbial biomass (Table 6.1). Disruption techniques, such as sonication or treatment with the enzyme lysozyme, are usually confined to laboratory-scale operations, due either to equipment limitations or on economic grounds. 

Highly-efficient general acid catalysis of acetal hydrolysis is involved in the reactions of glycosidase enzymes such as lysozyme (Dunn and Bruice, 1973)... 

In an earlier experiment, Jori et al. (14) reported that methionyl residues are important in maintaining the tertiary structure of lysozyme. The introduction of a polar center into the aliphatic side chain of methionine, as a consequence of the conversion of the thioether function to the sulfoxide, may bring about a structural change of the lysozyme molecule which, in turn, reduces the catalytic efficiency. When ozonized lysozyme was treated with 2-mercaptoethanol in an aqueous solution according to the procedure of Jori e al. (14), the enzyme did not show any increase in its activity. This may be explained in two ways. In one, such reactions are complicated by many side reactions, e.g. sulfhydryl-disulfide interchange, aggregation and precipitation of the modified enzyme (24-26). In the other, the failure to recover the activity of the enzyme may by associated with the extensive oxidation of other residues. 

Many bacterial enzymes and proteins, which are modified by the introduction of F-Phe or F-Tryp, have been obtained. Mammalian proteins containing F-Pro or other fluorinated amino acids have also been obtained, either in a direct manner in vivo) or, more efficiently, by the expression of the gene in a bacteria. Thus, trifluoro-methionine has been incorporated by E. coli in the lysozyme of a bacteriophage. Because this enzyme contains three methionines, it has been used to study the interactions of this protein with its ligands by F NMR. ... 

In order to understand why enzymes are such efficient catalysts, it is necessary to understand first why uncatalyzed reactions in solution are so slow. As illustrations, we consider the reactions that may be catalyzed by chymotrypsin or lysozyme. 

General acid-base catalysis is often the controlling factor in many mechanisms and acts via highly efficient and sometimes intricate proton transfers. Whereas log K versus pH profiles for conventional acid-base catalyzed chemical processes pass through a minimum around pH 7.0, this pH value for enzyme reactions is often the maximum. In enzymes, the transition metal ion Zn2+ usually displays the classic role of a Lewis acid, however, metal-free examples such as lysozyme are known too. Good examples of acid-base catalysis are the mechanisms of carbonic anhydrase II and both heme- and vanadium-containing haloperoxidase. 

Competitive Inhibition Enzyme Assays. Estimates of antibody-lysozyme dissociation constants can be obtained by taking advantage of the fact that most monoclonal antibodies efficiently inhibit enzymatic activity.3 5 The combining site of HyHEL-10, which is presented as an example, also has been demonstrated by X-ray crystallography to overlap a portion of the catalytic site of lysozyme.7 A constant concentration of lysozyme is incubated with varying amounts of antibody, and amounts of free (unbound) lysozyme molecules are estimated by the proportion of catalytic activity remaining. The assay assumes that the addition of Micrococcus lysodeikticus cell walls and concurrent dilution of the antibody-antigen mixture do not disturb the equilibrium. 

Figure 8 X-ray diffraction images, (a) A precession photograph of muconate lactonizing enzyme. The fourfold symmetry in the diffraction pattern is clearly visible. This gives an undistorted view of the reciprocal lattice but are no longer used because they are not as efficient as rotation images, (b) A rotation image of hen s egg white lysozyme. This easily obtainable image gives a distorted projection of the reciprocal lattice, but this is no obstacle for modern programs.

Part of the catalytic efficiency of lysozyme results from its ability to provide different solvent environments at the active site. This allows one catalytic group to exist in its acidic form at the same surrounding pH at which a second catalytic group exists in its basic form. This property is unique to enzymes chemists carmot provide different solvent environments for different parts of nonenzymatic systems. 

The values for chymotrypsin and acetylcholinesterase are within the range for normal metabolic enzymes. Lysozyme is an enzyme that degrades certain polysaccharide components of bacterial cell walls. It is present in many body tissues. Its low catalytic efficiency indicates that it operates well enough to catalyze polysaccharide degradation under normal conditions. 

The exo-p-D-2-acetamido-3-0-(D-l-carboxyethyl)-2-deoxyglucanase exo- -N-acetylmuramidase) produced by Bacillus subtilis growing under defined conditions is partly secreted and partly bound to the cell wall. The synthesis of this enzyme was induced, in decreasing order of efficiency, by a lysozymal digest of Micrococcus lysodeikticus cell walls, 2-acetamido-4-0-(2-acetamido-2-deoxy-P-D-gIuco-pyranosyl)-3-0-(D-l-carboxyethyl)-2-deoxy-D-glucopyranose, and 2-acetamido-4-0-[2-acetamido- 3-0-(d- 1 -carboxyethyl)-2-deoxy- P-D-glucopyranosyl]-2-deox y-D-... 

The yields of enzyme synthesized in vitro per ml of cell-free system were calculated from the specific activities of the purified enzymes (Zillig et al., 1966 Chamberlin et al., 1970). It was assumed that T7 lysozyme has the same specific activity as T4 lysozyme. The data on T4 enzyme yields had been described elsewhere (Schweiger and Gold, 1970). In the same cell-free mixtures, the amount of total protein synthesized was determined by the incorporation of C-leucine into TCA precipitable material. To the amount of total protein the yields of various individual enzymes were related (column 2). In order to estimate the percentage of senseful protein synthesized, we calculated the portion of a gene of the total stretch of DNA transcribed (28-30% in T7 40% in T4). The data in column 2 were divided by this factor, assuming that all proteins were synthesized with equal efficiency. 

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