Relation between stabilization and rigidification of the three-dimensional structure of an enzyme.
Purified cellobiase was coupled to periodate-oxidized dextran by reductive alkylation using sodium cyanoborohydride, sodium borohydride, and dimethylaminoborane for various reaction times. The thermal stability of the different conjugates obtained was studied and correlated to the number of links introduced between the enzyme and the soluble support. We observe that resistance to heat inactivation increases as a function of the number of modified lysines. Sodium cyanoborohydride was the most effective reducing agent. After 24 h reaction, the modification of 92% of the lysines gave a cellobiase-dextran conjugate that is a most stable enzyme. We conclude that the thermal stability observed for the chemically modified enzyme results from the rigidification of the three-dimensional structure of the protein. This rigidification increases with the number of links introduced between the enzyme and the polysaccharide. We also observe that chemical modification leads to a heterogeneous population of stabilized enzymes. Because of this heterogeneous population, it is necessary to develop a mathematical model of the kinetics of enzyme inactivation.
Germain
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Unité de Biochimie, Université Catholique de Louvain, Place Louis Pasteur, 1, B-1348 Louvain-La-Neuve, Belgium.
Slagmolen
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T
Crichton
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