Pectin, enzymic methylation

Three types of carbon-oxygen linkages may occur in a polyuronide ester, ether and glycoside. The ester-linked methyl group of pectins is split by cold dilute alkali. It may also be removed by certain enzymes. Methyl groups which often occur ether-linked to the uronic acid especially in plant gums and hemicelluloses, resist hydrolysis by mineral acid solutions of 2 to 4% concentrations, even in the autoclave. Such groups survive hydrolytic conditions that are usually chosen. 

Although the list of transmethylation reactions in which S-adenosylmethionine can function as the methyl donor could now undoubtedly be greatly expanded, the more recently added reactions, with the exception of the carbon methylations discussed below (Section II,C,6), appear from the chemical point of view to be further examples of types of reactions which were already known. Therefore, no complete compilation will be attempted. From the physiological standpoint, on the other hand, some of the reactions recently studied may be of great interest. For example, systems have now been reported for the enzymic methylation of RNA (Srinavasan and Borek, 1964 Rodeh et al, 1967), DNA (Oda and Marmur, 1966 Kalousek and Morris, 1969), pectin (Kauss and Hassid, 1967), and protein (Comb et al, 1966 Paik and Kim, 1968 Liss et al, 1969). The specific modifications of the properties of these biopolymers consequent to methylation may, of course, be important in... 

Kauss, H. and Swanson, A.L. (1969) Cooperation of enzymes responsible for polymerization and methylation in pectin biosynthesis. ZJ aturforsch. 24 28-33. 

Most cell walls are able to deesteriJfy exogeneous pectin very rapidly but in vivo these cell walls still contain large amounts of methyl-esterified pectins. How can the substrate and the enzymes coexist inside cell walls ... 

The possibility that the initial degree of methyl-esterification might be controlled by the properties of the methyltransferase enzymes was examined partial characterisation of these enzymes in suspension-cultured cells of fiax. Pectin methyltransferases beii enzymes characteristic of the Golgi apparatus [22], microsomes were fiactionated daily for ten days from suspension-cultured flax cells and incubated in the presence of C-SAM, the universal donor of methyl groups. 

Digestion of PGA by the PelL enzyme yielded a mixture of unsaturated ohgogalacturonides, giving evidence that PelL is an endo-deaving lyase (17). An exo-enz3mie, such as the EC 16 PelX, would generate a single product (15). The PelL protein differs from the major E. chrysanthemi pectate lyases in its ability to cleave both PGA and methylated pectin (17). The PelL activity has a basic optimum pH and an absolute requirement for Ca + ions. Analysis of culture supernatants demonstrated that PelL is an extracellular enzyme, such as the other secondary pectate lyases (17). 

Each fruit has specific quantities and ratio of pectin, hemicelluloses and cellulose. These polysaccharides are important concerning enzymes activities required to produce juices and concentrates. Moreover, even if molecular weight and methylation degree of the pectin are specific for each fruit, during the fruit maturation, endogenous pectinases depolymerases and esterase are changing the pectin characteristics This broad variability of raw material makes difficult the standardisation of fruits processing. 

With the use of exogenous pectinmethylesterase, the methanol content is lower because the demethylation of the pectin is not complete, part of the methylated pectin remaining in the coagulum and because the contact time between the PME and the pectin is shorter than in the past with the natural enzyme. 

The chromatogram of the serum from juice with poor cloud stability obtained after mash treatment with the enzyme mixture including the pectin depolymerisation enzyme combination of polygalacturonase and pectin methyl esterase is seen to be lacking the A-peak. High molecular pectin is therefore considered to be a prerequisite for a cloud stable juice. 

Figure 1 indicates that pectin methyltransferase (PMT) activity from freeze-thawed microsomes measured without exogenous substrate was maximal at neutral pH (6.5 to 7.5). When exogenous pectic substrates of various DE had been added, similar optimal neutral pH was observed, and the activity was slightly stimulated (1.2 to 1.8 times). A second optimal pH occured at pH 5.5, but in the presence of low methylated pectin (DE 0.1). As suggested by Lineweaver and Ballou [8] to explain the behaviour of another pectic enzyme -i.e. pectin methylesterase (PME), the mobility and the activity of PMT might be influenced by the presence of polyanionic substrates. On the other hand, the existence of several forms of pectin methyltransferase in flax microsomes might be responsible for such variations of the activity. 

Enzymes can be used to specifically modify the pectins. Pectin methyl esterase is already widely used to adjust the gelling properties of commercially available pectins. The acetyl esters also strongly affect the gelation [2,3] and removal is important for the upgrading of sugar beet pectin, extractable from a by-product of the sugar industry. 

We described here the characterisation of the pemB gene and its product the second PME of E. chrysanthemi. The biochemical analysis of the purified protein indicated that PemB is actually an enzyme that demethylates pectin, leading to formation of methanol and PGA. However, PemB is more active on methylated oligogalacturonides than on polymeric pectin. The activating effect of non-ionic detergents on PemB was never pointed out for other pectinases and it is a characteristic of many membrane enzymes (21). 

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