Maltose acceptor products

M. Dols, M. Remaud-Simeon, R. M. Willemot, M. Vignon, and P. Monsan, Structural characterization of the maltose acceptor products synthesized by Leuconostoc mesenteroides NRRL B-1299 dextransucrase, Carbohydr. Res., 305 (1998) 549-559. 

It was concluded from the study that a high yield of four to six maltose acceptor products and a low yield of dextran could be obtained at any 1 1 maltose-to-su-crose concentration by using a high concentration of enzyme. A single acceptor product (panose in the case of maltose as an acceptor) may be obtained free of dextran by using a concentration of 100 mM sucrose at a maltose-to-sucrose ratio of 20 1 and an intermediate concentration of enzyme of 1 U/mL. 

The final product concentration depends significantly on the ratio of substrate and acceptor. The first acceptor product (panose in the presence of maltose) is favored at high maltose in excess, further tetra- and pentasaccharides are formed in significant amounts.37 Thus isomalto-oligosaccharides (IMOS) can be produced in good yield under appropriate reaction conditions (see Section III). 

Summarizing the results of many investigations, monosaccharides and such derivatives as D-mannitol and D-glucitol are rather weak acceptors. Disaccharides, including such acceptor products as isomaltose, are much better acceptors, except for certain molecules, for instance leucrose, which is not an acceptor.29,46,47 The decrease of enzyme activity with time has been described in terms of a first-order reaction. The inactivation parameters have been calculated for the immobilized enzyme. The inactivation constants kd were 0.0135 (1/d) when maltose was the acceptor (stabilizing), and 0.029 (1/d) when fructose was the acceptor.38... 

Using a constant 1 1 ratio of maltose to sucrose and different concentrations from 1.25 to 200 mAf, the concentration of enzyme was varied 1000-fold from 0.12 to 120 U/mL. As the concentration of enzyme was increased, the amount of dextran formed decreased and the amount of acceptor products formed increased. The decrease in dextran was most pronounced at the lower substrate concentrations. At the highest enzyme concentration (120 U/mL), all of the substrate concentrations (1.25 to 200 mAf) gave just 5% dextran out of the total amount of product formed. 

Even though the reaction scheme is complex, attempts have been made to model the reaction and develop kinetic equations. Mooser et al.lu developed kinetic equations for a limited case based on the primer mechanism. Reh et a/.145 developed kinetic equations for the synthesis of dextran and for the maltose acceptor reaction based on the two-site insertion mechanism. Likewise, Boker et al,146 developed a kinetic equation for the formation of the fructose acceptor product, leucrose, based on the two-site insertion mechanism that included an acceptor site. An equation was derived for the overall reaction rate that was a function of the consumption of sucrose by dextran formation and acceptor-product formation. 

Isomaltooligosaccharides (as mixtures of different a-glucooligosaccharides) are produced in part by dextransucrases using sucrose as the substrate and maltose as an acceptor. They have been used as sweeteners in Europe and Japan for years, mainly in the field of prebiotics, and dermocosmetics [11-13]. Several different methods can be used to produce them by using such carbohydrates as sucrose, maltose, starch, and dextran. For the industrial production of a-glucooligosaccharides with sucrose as the substrate and maltose as the acceptor by one specific dextransucrase (of L. mesenteroides NRRL B-1299), the kinetic behavior of the enzyme has been characterized in order to optimize the synthesis [14]. 

To try to determine the number of subsites of the acceptor part of the active site we incubated 1,6-anhydro-maltose 13 and maltotriose 14 with CGTase in the presence of a-CD. Only compound 14 gave coupling products at a reasonable rate. It is suggested that two glucosyl residues are recognized on a coupling reaction so only two subsites constitute the acceptor part of the active site. 

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