P55: Improvement of the stability of raw starch digesting amylase (RSDA) from Aspergillus carbonarius by succinylation

Raw starch digesting amylases (RSDAs) have enormous potentials in biotechnology limited by enzyme instability. To improve the stability of RSDA from Aspergillus carbonarius, RSDA was chemically modified by varying concentrations in mg (10, 15 and 20) of succinic anhydride. The catalytic activities and thermostabilities were compared. Succinylation levels were determined using trinitrobenzene sulphonic acid (TNBS) assay. Circular dichroism and fluorescent intensity spectra were used to probe structural changes to the modified enzyme. Modification with led to a slight loss of enzyme activity depending on the concentration used; while 20 mg SA caused 20% loss of activity, 10 mg SA resulted in only 6 % loss of native enzyme activity. The pH optimum of 5 was altered to pH 3 using 10 mg SA and pH 4 when 15 mg or 20 mg SA was used for modification. Modification remarkably improved pH stability of enzyme especially in acidic conditions. At pH 3 the 10 mg SA derivative lost only 10% of its activity compared to the native enzyme which lost over 42% of its activity. Optimum temperature shifted from 30oC to 40oC for 10 mg and 15 mg SA derivatives while modification with 20 mg SA did not alter the temperature optimum. Modified enzyme retained 100 % of its activity after 20 min incubation at 80oC. Using 10 mg SA to modify the lysine groups of the RSDA proved a simple and effective way of improving the stability profile of the RSDA.