Preparation of the stabilized glycoenzymes by cross-linking their carbohydrate chains |
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Authors: | Branko Kozuili? Ivan?ica Leu?tek Branka Pavlovi? Pavao Mildner Slobodan Barbari? |
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Institution: | 1. Laboratory of Biochemistry, Faculty of Food Technology and Biotechnology, University of Zagreb, Pierottijeva 6, 41000, Zagreb, Yugoslavia
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Abstract: | Each of the three high-mannose type glycoproteins studied, acid phosphatase, invertase, and glucose oxidase, could be specifically
cross-linked through its carbohydrate chains. The procedure involves periodate oxidation of carbohydrate residues followed
by reaction of the generated aldehyde groups with adipic acid dihydrazide as a cross-linker. The amount and size as well as
solubility of the formed polymers could be efficiently controlled by varying the reaction conditions, i.e., the oxidation
degree and the concentrations of glycoproteins, cross-linker, and hydrogen ions during the cross-linking reaction. It was
found that the quantity and size of polymers increased with oxidation degree and protein concentration and by lowering the
pH. When the protein concentration was above and pH below certain values, depending on the glycoenzyme, insoluble polymers
formed. The soluble cross-linked polymers retained a high level of original activity, and the minor decrease in specific activity
noticed was shown to occur during the periodate oxidation step. The cross-linked glycoenzymes are much more resistant to denaturation
by high temperature and by changes in pH, demonstrating the usefulness of this method in preparation of the stabilized glycoprotein
derivatives. |
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Keywords: | Glycoproteins cross-linking stabilization glycoenzymes quarternary structure |
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