Pectolytic enzyme

Glycosidases (cellulases and amylases from As-pergillus niger) in combination with proteinases are recommended for removal of shells from shrimp. The shells are loosened and then washed off in a stream of water. 

The cell walls of gram-positive bacteria are formed from peptidoglycan (synonymous with murein). Peptidoglycan consists of repeating units of the disaccharide N-acetylglucosamine 

Proteins from seeds of the mustard family (Brassicaceae), such as turnip, rapeseed or brown or black mustard, contain glucosinolates which can be enzymatically decomposed into pungent mustard oils (esters of isothiocyanic acid, R—N=C=S). The oils are usually isolated by steam distillation. The reactions of thiogly-cosidase and a few glucosinolates occurring in Brassicaceae are covered in section 17.1.2.6.5. 

Pectolytic enzymes are described in section 4.4.5.2. Pectic acid which is liberated by pectin methylesterases flocculates in the presence of Ca + ions. This reaction is responsible for the undesired cloud flocculation in citrus juices. After thermal inactivation of the enzyme at about 90 °C, this reaction is not observable. However, such treatment brings about deterioration of the aroma of the juice. Investigations of the pectin esterase of orange peel have shown that the enzyme activity is affected by competitive inhibitors oligogalacturonic acid and pectic acid (cf. Fig. 2.51). Thus, the increase in turbidity of citrus juice can be prevented by the addition of such cort5)ounds. 

Pectinolytic enzymes are used for the clarification of fruit and vegetable juices. The mechanism of clarification is as follows the core of the turbidity causing particles consists of carbohydrates and proteins (35%). The prototropic groups of these proteins have a positive charge at the pH of fruit 

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Subsequently, a clear Juice is obtained by ultrafiltration. A serious problem in this process is the fouling of the ultrafiltration membrane, causing a reduced flux rate. For apple processing, the material responsible for this effect has been isolated and extensively characterized [2-4]. It appeared to consist mainly of ramified pectic hairy regions (MHR), which were not degraded by the pectolytic enzymes present in the technical pectinase preparation. 

Pectins and pectolytic enzymes in relation to development and processing of green beans (Phaseolus vulgaris L.). 

Aim of this work was to optimise enzymatic depolymerization of pectins to valuable oligomers using commercial mixtures of pectolytic enzymes. Results of experiments in continuous and batch reactor configurations are presented which give some preliminary indications helpful to process optimisation. The use of continuous reactors equipped with ultrafiltration membranes, which assure removal of the reaction products, allows to identify possible operation policy for the improvement of the reaction yield. 

The catalytic capacity of several excreting pectolytic enzymes obtained from various yeast strains was examined using in vivo and biochemical techniques. Of the 33 yeast strains studied 30 were isolated from champagne wine during alcoholic fermentation. Only one yeast strain was found to excrete pectolytic enzymes and was identified as Saccharomyces cerevisiae designated SCPP. Three types of pectolytic enzymes were found to be excreted by SCPP polygalacturonase (PG), pectin-lyase (PL) and pectin-esterase (PE) [1]. 

These results prompted us to examine the characteristics of the extracellular pectolytic enzymes secreted in medium supplemented with glucose and galactose. Figure 2 shows the profile of elution of pectolytic activities recovered from the flow-through of a DEAE-cellulose column chromatographed on a CMC-cellulose column. 

Figure 2. CM-cellulose chromatography of pectolytic enzymes. The activity peaks of the flow-through of a DEAE-cellulose chromatography was applied to a CM-cellulose column. The column was eluted with a NaCl (0-0.5M) continuous gradient at a flow rate of 34 ml/h. 10 ml fractions were collected and assayed for pectolytic activities Symbols (0) pectate lyase ( ) polygalacturonase (reducing sugar-releasing activity) (x) protein. Other details in Methods.

Table 1 shows some biochemical properties of the pectolytic enzymes present in pool 1. The pectin lyase/pectate lyase activities (pool I) and polygalacturonase activity (pool II) were not significantly affected by NH4+, Na+ and K+ (0,25 - 2,5mM), while Al +, p-mercaptoethanol, Hg2+, EDTA, Ba + and Zn+2 (2,5mM) inhibited 30-100% these activities. On the other hand, Ca2+, Mg + and Mn + at 2,5mM concentration activated 20-100% pectin/pectate lyases but Ca " " and Cu " " (2,5mM) inhibited polygalacturonase activity about 42 - 70%. 

Only PAE, a novel acetyl esterase, could remove acetyl from beet pectin, to a maximum of 30%. This was shown to be one specific acetyl group in theJiomogalacturonan chain of pectin (smooth region) by NMR spectroscopy. PAE activity was influenced by buffer salts and the addition of bivalent cations. PAE worked cooperatively with pectolytic enzymes. 

FAE was identical to these enzymes in requiring an endoxylanase for activity on xylans. Under these circumstances ferulic acid could be released too. It has been observed that for the complete breakdown of different xylans, different enzyme combinations are required, as the substitutions on the xylan chain can vary widely [7]. No activity could be shown on pectins, neither in combination with other AE s and/or pectolytic enzymes. 

After purification of the enzyme preparation both at Sephadexes of various types and at Toyopearl HW—56 two fractions were obtained (fractions 1 — 2). One of them (fraction 1) possessed pectinesterase activity under the condition of absence of the activity of other pectolytic enzymes. ... 

Since many years, pectolytic enzymes have been widely used in industrial beverage processing to improve either the quality and the yields in fruit juice extraction or the characteristics of the final product [1,2]. To this purpose, complex enzymatic mixtures, containing several pectolytic enzymes and often also cellulose, hemicellulose and ligninolytic activities, are usually employed in the free form. The interactions among enzymes, substrates and other components of fruit juice make the system very difficult to be investigated and only few publications are devoted to the study of enzymatic pools [3-5], An effective alternative way to carry out the depectinisation process is represented by the use of immobilized enzymes. This approach allows for a facile and efficient enzymatic reaction control to be achieved. In fact, it is possible to avoid or at least to reduce the level of extraneous substances originating from the raw pectinases in the final product. In addition, continuous processes can be set up. 

Enzyme-mediated action, by sulfhydryl and phenylalanine-lyase (PAL) enzymes as well as by other enzymes such as cellulase, catalase, peroxidase, phosphorylase and pectolytic enzymes ... 

Pectinases and (3-glucanases are the only enzymes allowed in wine-making by European legislation. They are used as clarification and filtration agents and also to release aroma compounds that are mostly present in grape as nonvolatile glycosidic precursors. Pectolytic enzymes are also reported to increase extraction of phenolic compounds and wine color... 

Femandez-Zurbano, P. et al.. Effects of maceration time and pectolytic enzymes added during maceration on the phenolic composition of must. J. Food Sci. Technol. Int. 5, 319, 1999. 

Treatment of pulp with pectolytic enzymes has two additional benefits. Free-run juice is released more readily from the pulp and overall juice yield is increased. Studies with heated Concord pulp containing cellulose fibers have shown that from 1000-gram portions of pulp, free-run juice obtained 5 min after placing the pulp on an 8-mesh screen was increased from 337 ml for controls with no enzyme to an average of 387 ml for lots containing pectolytic enzymes (at a concentration equivalent to that used in commercial operations). This represented an increase of 15% in free-run juice. Total yield, combining both free-run and hand-pressed juice, increased by 3%. (Equivalent yield on a commercial basis would have been 193.6 gal/ton.)... 

In cold pressing pulp for white wines, present practices vary considerably within the industry. Some processors using continuous presses have found it an advantage to treat pulp with pectolytic enzymes prior to pressing. However, with the Willmes press, cold pulp to which cellulose fibers have been added press with no difficulty, and juice yields have been considered satisfactory. A general practice has been to add pectolytic enzymes to white must before yeast inoculation as an aid to rapid clarification of the new wine. 

If juice is to be sold as not from concentrate it is usually screened and pasteurised immediately after pressing - an operation with two main objectives. The first is to control the growth of spoilage micro-organisms that live on the fruit surface (mainly yeasts and moulds). The second is to destroy the pectolytic enzymes that occur naturally in fruit that would otherwise break down the cloudy nature of the juice. If, however, a clear juice is required (e.g. apple or raspberry), enzymes can be added to accelerate this natural process. 

The ability of growth substances to modify the activity of pectolytic enzymes appears to have great ecological significance. This is particularly true for allelopathic compounds. 

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