Glucan-Active Enzymes

Table 4.6.14 Enzyme activities (1,4-a-glucan branching enzyme) in liver, muscle, and fibroblasts... 

Enzymatic hydrolysis, of p-glucan, 753 Enzyme activity measurements conditions, activity units, and reaction rate, 331 -334... 

Another purified enzyme preparation which produces laminaripentaose from insoluble laminarin and from heat-treated pachyman is produced by a strain of Arthrobacter luteus (100,101,102) when grown on yeast cells or / -(1 —>3)-glucan. The enzyme, which was named Zymolase (also referred to as Zymolyase) appeared to be homogeneous by electrophoresis in a Tiselius apparatus and by ultracentrifugation. The molecular weight of the enzyme was estimated from ultracentrifugation to be ca. 20,500. The optimum pH for lysis of viable yeast cells was 7.5. The optimum temperature was 35°C. The optimum pH for heat-treated pachyman hydrolysis was 6.5, and the optimum temperature was 45°C. A Lineweaver-Burk plot with heat-treated pachyman yielded a Km value of 0.04% when the solubilized carbohydrate was assayed by the phenol-sulfuric acid method. Zymolase lost all its activity after incubation at 60°C for 5 min. 

In higher plants several glucan metabolizing enzymes occur as plastid- and cytosol-specific isozymes (1,2). The dual intracellular location of these enzyme activities suggests that both the plastidic and the cytosolic compartment contain a pool of polysaccharides. However, until now a cytosolic starch-like polysaccharide which functions as the physiological carbohydrate substrate of the cytosol-specific isozymes has not been identified and, therefore, the metabolic function of these enzyme forms remains enigmatic. 

Cline, K. and P. Albersheim, Host pathogen interactions. XVII. Hydrolysis of biologically active fungal glucans by enzymes isolated from soybean cells. Plant Physiol., 65,221-228 (1981). 

Aloni Y, Delmer D.P., and Benziman M. 1982. Achievement of high rates of in vitro synthesis of 1, 4-beta-D-glucan activation by cooperative interaction of the Acetobacter xylinum enzyme system with GTP, polyethylene glycol, and a protein factor. Proc Natl Acad Sci USA 79(21) 6448-6452. Amikam D. and Benziman M. 1989. Cyclic diguanylic acid and cellulose synthesis in Agrobacterium tumefaciens. J Bacterid 171(12) 6649-6655. 

An ejco-/3-l,4-D-glucanase (eA u-j8-l,4-D-glucosidase) isolated from almond seeds acted on -D-glucans and jS-D-gluco-oligosaccharides by a mechanism similar to that of glucoamylase. A multiple (two)-subsite model was suggested for the active enzyme-carbohydrate complex on the basis of substrate specificities and rate data. 

Fig. 2.2 Simplified schema of the prophenoloxidase activating (melanisation) cascade. Pattern recognition protein will, after binding to compounds (e.g. lipopolysaccharide, 3-l,3-glucans or peptidoglycans) typical for microorganisms, trigger activation of upstream components of serine proteinase cascade.The terminal proteinase will cleave prophenoloxidase to the catalytically active enzyme phenoloxidase. Different phenolic substances will give rise to melanin as well as toxic short-lived intermediates.

Various strains of oral streptococci produce D-glucosyltransferases which utilize sucrose as a o-glucosyl donor in the production of soluble and insoluble D-glucans. Consequently, it may be expected that some deoxyfluoro derivatives of sucrose function as competitive inhibitors for the dextransu-crases of tooth bacteria, thus preventing decay, or at least may be used as active-site probes for the enzymes. Another aim of these researches is to find non-metabolizable sweeteners. 

The echinocandins have a unique target for their antifungal activity—specifically, (5-1,3-glucan synthase, an enzyme that produces an important component of the fungal wall. Caspofungin is currently the only product of this class that is FDA approved it is indicated for treatment of IA refractory... 

By rect comparison of structural and functional characteristics of different, naturally-occurring enzymes and modified biocatalysts, researchers hope to delineate the structural features controlling the hydrolytic and S3mthetic catalytic activities of this family of glucosylases. This understanchng will eventually lead to the ability to "engineer" glucan biocatalyst capability. 

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