Characterization of free and immobilized invertase regarding activity and energy of activation

Abstract

Invertase from NOVO Nordisk has been immobilized in controlled pore silica particles (diameter: 0.351 mm and mean pore size: 37.5 nm) by covalent binding with the silane-glutaraldehyde method. The activity of the free and immobilized enzyme (IE) was determined with 5% (w/v) sucrose, at 35 to 65ºC and pH from 3 to 7. Maximum activities were found in the pH range from 5 to 6 for free invertase, and pH 4.5 for the IE. Activity yield for the IE was 24%. The Energy of Activation (Ea) was found to be a function of pH, giving for free invertase, Ea = 7.0 and 6.86 kcal/mol at pH 5.0 and 5.5, respectively, whereas for the immobilized enzyme, Ea = 6.55 and 5.93 kcal/mol at pH 4.5 and 5.0, respectively

Immobilization of invertase on celite and on polyacrylamide by an absorption procedure

“…Similar pH values were reported by Godbole et al 4 and Arruda and Vitolo, 2 who found that both soluble and immobilized invertase activity presented an optimum at pH 4.6. However, these results differ from those noted by Bergamasco et al, 5 who noted that the immobilization of invertase in controlled silica particles by the covalent binding with the silane-glutaraldehyde shifts the optimum pH of the enzyme by about 0.5 unit from 5.0 to 4.5. Sun et al 13 reported that the carrier with charge would induce the optimum pH of the immobilized enzyme to change: to higher values for a carrier with a cationic charger and lower values for a carrier with an anionic charger.…”

Immobilization of the recombinant invertase INVB from Zymomonas mobilis on Nylon-6

“…2). These results indicate that immobilization on Nylon-6 happens more quickly than on other supports [15], and is similar to that for the fusion protein INVB-CBD cex , when immobilized on crystalline cellulose [16].…”

“…Other reports [15,[18][19][20] show optimal pH for immobilized invertases to be in the range of 3.0-6.1. Previous studies have indicated that both the support and the method used for immobilization are the factors, which most influence the optimum pH [15].…”

“…These data suggest that the kinetic parameters of re-INVB were affected, when immobilized on Nylon-6, probably due to steric hindrance between the immobilized enzyme and the insoluble support [23]. A decline in catalytic activity for the invertase and other glycoenzymes upon immobilization has been reported [15,24,25]. Immobilization of invertase of yeast in poly(maleic anhydride-hexen-1) membrane [26] or in poly(vinyl alcohol) membrane suggested that the formation of the enzyme-substrate complex was more difficult for immobilized invertase, than for native invertase [27].…”

Sucrose hydrolysis catalyzed by auto-immobilized invertase into intact cells of Cladosporium cladosporioides

“…These results show different pH values for each invertase, for different microorganisms that produce it. The temperature that optimizes the hydrolysis process is a function of the enzyme type and experimental conditions; values within 45 to 73ºC interval have been reported (Bergamasco et al 2000;Oda and Tonomura, 1994).…”