COMPARATIVE STUDY ON KINETIC PROPERTIES OF FREE ANDIMMOBILIZED ASPARAGINASE FROM LOCAL ISOLATE OFPseudomonase aeruginosa

Abstract

The purified asparaginase from local isolate of P. aeruginosa.was immobilized
by gel entrapment using polyacrylamide,agar,sodium alginate,ionic binding on
ionexchanger using DEAE-cellulose and immobilized in gelatin by cross-linking
with glutaraldehyde.
The later method was the most efficient method for immobilization since the
immobilized enzyme retained about 53.5% of original activity of free enzyme. The
optimal pH for immobilized enzyme activity was 7.5 and the enzyme was most
stable at pH 6-8.5, while the values of optimal pH for activity and Stability of free
enzyme were 8.5 and (7.5-8) respectively. The immobilized enzyme retained its
original activity when incubated at 25-45C° for 30 min., while the free enzyme
retained its original activity after 30 min. at 25-37C°. The obtained values of
Michaelis constant (Km) and Vmax for immobilized enzyme were 0.46×10-4 M and
12.2 μM/min.,respectively ,and the Km and Vmax for free enzyme were 0.25×10-4
M and 16.7 μM/min,respectively. The immobilized enzyme was highly specific
towards L-asparagine and L-glutamine. The free and immobilized enzyme retained
about 90.2% and 51% of its original activity after 3 weeks of storage at 4 C°.