Lysozyme catalyzed reactions

Baneq ee, S.K., Kregar, I., Turk, V., Rupley, J.A. Lysozyme-catalyzed reaction of the N-acetylglucosamine hexasaccharide. Dependence of rate on pH. J. Biol. Ghem. 1973, 248,4786-92. 

Fukamizo, T., Minematsu, T., Yanase, Y, Hayashi, K., and Goto S. 1986a. Substrate size dependence of lysozyme-catalyzed reaction. Arch Biochem. Biophys. 250 312-321. 

Kuhara, S., Ezaki, E., Fukamizo, T., and Hayashi, K. 1982. Estimation of the free energy change of substrate binding lysozyme-catalyzed reactions. J. Biochem. 92 121-127. 

Masaki, A., Fukamizo, T., Otakara, A., Torikata, T., Hayashi, K., and Imoto, T. 1981a. Lysozyme-catalyzed reaction of chitooligosaccharides. J. Biochem. 90 527-533. 

Natural catalysts such as ribonuclease A and lysozyme catalyze reactions by using both GA and GB catalysis through transition states TS27 and TS28, respectively, during the course of catalytic reactions. Whether the occurrence of GA and GB catalyses in ribonuclease A-catalyzed reactions is concurrent (TS27) or stepwise is still a topic of debate. But the essence of catalysis, i.e., involvement of both GA and GB catalyses is almost certain. 

Thus, investigation of the kinetics of lysozyme-catalyzed lysis of bacterial cell walls has shown that the presence of an organic solvent decreased the overall reaction rate reversibly with the rados of the maximum rate constant of lysis in water and in mixed solvent given by... 

Observations carried out on lysozyme in mixed solvents as a function of temperature demonstrate that lysozyme-catalyzed lysis can he performed under such abnormal conditions and that the reaction can be quenched at subzero temperatures and resumed by heating. The problem is now to check whether one can obtain an enzyme-substrate complex (in this case lysozyme-oligosaccharide) stabilized at low temperature. Such a complex can be detected by differential absorption spectroscopy. Difference spectra in the UV region (240-320 nm) were recorded. The reference cell contained a solution of lysozyme (1.39 x 10 M) and the sample cell contained a solution of lysozyme at the same concentration plus substrate, here hexa-NAG (1.66 x lO " M). The buffer used is acetate at pH 5 plus organic solvent. Also, difference spectra have been determined in the presence of the small-chain sac-... 

Lysozyme catalyzes the hydrolysis of the polysaccharide component of plant cell walls and synthetic polymers of j8(l — 4)-linked units of A-acetylglucosamine (NAG) (Chapter 1). It is expected from studies on nonenzymatic reactions that one of the intermediates in the hydrolytic reaction is a oxocarbenium ion in which the conformation of the glucopyranose ring changes from a full-chair to a sofa (half-chair) conformation (Chapter 1). The transition state analogue I, in which the lactone ring mimics the carbonium ion-like transition state n, binds tightly to lysozyme = 8.3 X 10 8M.10... 

Fig. 6. Proposed reaction mechanisms by which lysozyme catalyzes the cleavage of a polysaccharide substrate. Scheme I is that proposed by Post and Karplus (1986) and indicates the possibility of cleavage with no prior assumption of distortion of the sugar ring at site D. Scheme II is that originally developed by Blake, Johnson, Phillips, and co-workers and involves ring distortion as a critical step. (Reproduced with permission from Post and Karplus, 1986.)...

Six classes of enzyme-catalyzed reaction are recognized in systematic nomenclature [61]. Their names and the type of chemical reaction catalyzed by each are indicated in Table 4.1. All enzymes have systematic names based on the above, but many are known by historically important trivial names e.g. trypsin, chymo-trypsin, pepsin, lysozyme, catalase. 

The rate of an enzyme-catalyzed reaction as a function of pH generally yields a bell-shaped curve. Some enzymes are very sensitive to small changes in pH (lysozyme), whereas others (POase) are relatively insensitive (within 1-2 pH units near their optima). Enzymes with similar activities but from different origin may have very different optima. APase from Escherichia coli is optimally active at a pH of about 8, whereas APase from calf intestine is most active around pH 10 and the activity of these enzymes decreases strongly outside their optima. Nevertheless, the E. coli enzyme is often assayed at the pH optimum of the intestinal enzyme. The optimum substrate concentrations may also be pH dependent for some enzymes (Chapter 10). 

Nicotinamide adenine dinucleotide (NAD) is the coenzyme form of the vitamin niacin. Most biochemical reactions require protein catalysts (enzymes). Some enzymes, lysozyme or trypsin, for example, catalyze reactions by themselves, but many require helper substances such as coenzymes, metal ions, and ribonucleic acid (RNA). Niacin is a component of two coenzymes NAD, and nicotinamide adenine dinucleotide phosphate (N/kDP). NAD (the oxidized form of the NAD coenzyme) is important in catabolism and in the production of metabolic energy. NADP (the oxidized form of NADP) is important in the biosynthesis of fats and sugars. 

The fi-glucan exo- and endo-hydrolases are discussed with reference to newer techniques for the investigation of their specificity and action pattern. Those exo-hydrolases which have been well characterized are described individually. The endo-hydrolases are examined from the point of view of their linkage specificity, action on substituted glucans and their specificity for various monomer units. The significance of more random and less random endo-action patterns is considered in relation to single or multiple attack mechanisms. Certain features of p-glucan endo-hydrolase catalyzed reactions are discussed in relation to current views on the three-dimensional structure and mechanism of action of lysozyme. 

Lysozyme has two catalytic groups at the active site Glu 35 and Asp 52 (Figure 24.9). Once it was determined that the enzyme-catalyzed reaction takes place with retention of configuration at the anomeric carbon, it could be concluded that it cannot be a one-step Sn2 reaction the reaction must involve at least two steps and, therefore, must form an intermediate. Although lysozyme was the first enzyme to have its mechanism studied— and the mechanism has been studied extensively for almost 40 years—only recently have data been obtained that support the mechanism shown in Figrue 24.9 ... 

Akiyama, K., Kawazu, K., and Kobayashi, A. 1995. A novel method for chemo-enzymatic synthesis of elicitor-active chitosan oligomers and partially Af-acetylated chitin ohgomers using M-acetylated chitotrioses as substrate in a lysozyme-catalyzed transglycosylation reaction system. Carbohydr. Res. 279 151-160. 

The Mechanism for an Enzyme-Catalyzed Reaction That Involves Two Sequential S j2 Reactions 1123 PROPOSED MECHANISM FOR THE REACTION CATALYZED BY LYSOZYME... 

Redox catalysts can be stabilized by encapsulation during silica sol-gel formation [33-35], in which the conductivity of the silica matrix is achieved by coimmobilization of a conductive material, such as carbon nanotubes (CNTs). The cationic protein lysozyme catalyzes and templates the formation of silica directly onto a conductive carbon paper electrode. Inclusion of CNT and glucose oxidase (GOx) into the reaction mixture results in a catalytic composite that becomes encapsulated as the silica forms [36]. 

Procarboxypeptidase A is activated by the removal of a peptide of some 64 residues from the N-terminus by trypsin.153 This zymogen has significant catalytic activity. As well as catalyzing the hydrolysis of small esters and peptides, procarboxypeptidase removes the C-terminal leucine from lysozyme only seven times more slowly than does carboxypeptidase. Also, the zymogen hydrolyzes Bz-Gly-L-Phe with kcsA = 3 s-1 and KM = 2.7 mM, compared with values of 120 s 1 and 1.9 mM for the reaction of the enzyme.154 In contrast to the situation in chymotrypsinogen, the binding site clearly pre-exists in procarboxypeptidase, and the catalytic apparatus must be nearly complete. 

In one case, a small peptide with enzyme-like capability has been claimed. On the basis of model building and conformation studies, the peptide Glu-Phe-Ala-Ala-Glu-Glu-Phe-Ala-Ser-Phe was synthesized in the hope that the carboxyl groups in the center of the model would act like the carboxyl groups in lysozyme 17). The kinetic data in this article come from assays of cell wall lysis of M. lysodeikticus, chitin hydrolysis, and dextran hydrolysis. All of these assays are turbidimetric. Although details of the assay procedures were not given, the final equilibrium positions are apparently different for the reaction catalyzed by lysozyme and the reaction catalyzed by the decapeptide. Similar peptide models for proteases were made on the basis of empirical rules for predicting polypeptide conformations. These materials had no amidase activity and esterase activity only slightly better than that of histidine 59, 60). 

Figure 4. Essential features of the hydrolytic reaction catalyzed by hen egg white lysozyme. Notice that two nearby carboxyl groups should be able to stabilize the oppositely charged oxocarbonium ion intermediate.

Kinetic isotope effect for lysozyme. A secondary kinetic isotope effect is expected because a molecule with H in the number 1 position can be converted to the corresponding oxocarbenium ion somewhat more easily than the molecule with 2H in the same position (Eq. 12-13). For example, in the nonenzymatic acid-catalyzed hydrolysis of a methyl- glucoside, a reaction also believed to proceed through a carbocation intermediate,41 75 the ratio h / h, is 1.14 for the a anorner and 1.09 for the (3 anorner.53 In the base-catalyzed hydrolysis of the same compound, which is believed to occur by a double-displacement reaction involving participation of the neighboring OH group on C-2, the ratio /c1h / /c2h is 1.03. The corresponding ratio measured... 

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. 

---