Alpha-amylase immobilized on plastic supports: stabilities, pH and temperature profiles and kinetic parameters.
The covalent immobilization of alpha-amylase on new isocyanate, acid chloride and carboxylic acid--activated plastic supports shows the viability of such supports for immobilizing enzymes, especially those reacting with 1,6-diaminohexane and glutaraldehyde for producing side arms. The operational stability of immobilized alpha-amylase could be extended by crosslinking the enzyme or by extending the support's side arm (substrate concentration has no effect). Inactive immobilized alpha-amylase were unfolded and then refolded at elevated temperature, these supports were found to be essential in increasing the stability of the enzyme during refolding. The pH curves for the immobilized enzyme were in general found not to be shifted from the soluble enzyme's pH optimum, although one isocyanate plastic support derivative shifted the pH activity profile of alpha-amylase to a higher range by 1.5 pH units, probably due to reaction between the enzyme and the free anhydride groups existing on the support's surface. In all cases, the immobilized enzyme's temperature activity profiles were shifted to a lower temperature range when compared to the soluble enzyme. The immobilized alpha-amylase Michaelis constants increased and the the maximum rates and specific activities decreased when compared to the soluble enzyme kinetic parameters.
Roig
M G
MG
Departamento de Química Físca, Facultad de Farmacia, Universidad de Salamanca, Spain.
Slade
A
A
Kennedy
J F
JF
D001412:Bacillus subtilis / Q000201:enzymology; D002264:Carboxylic Acids / Q000737:chemistry; D002712:Chlorides / Q000737:chemistry; D004793:Enzyme Reactivators; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000378:metabolism*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D011126:Polypropylenes / Q000737:chemistry*; D011489:Protein Denaturation; D012995:Solubility; D013861:Thiocyanates / Q000737:chemistry; D000516:alpha-Amylases / Q000378:metabolism*
D002264:Carboxylic Acids; D002712:Chlorides; D004793:Enzyme Reactivators; D004800:Enzymes, Immobilized; D011126:Polypropylenes; D013861:Thiocyanates; D000516:alpha-Amylases