Thermal inactivation and product inhibition of Aspergillus terreus CECT 2663 alpha-L-rhamnosidase and their role on hydrolysis of naringin solutions.
The kinetics of thermal inactivation of A. terreus alpha-rhamnosidase was studied using the substrate p-nitrophenyl alpha-L-rhamnoside between 50 degrees C and 70 degrees C. Up to 60 degrees C the inactivation of the purified enzyme was completely reversible, but samples of crude or partially purified enzyme showed partial reversibility. The presence of the product rhamnose, the substrate naringin, and other additives reduced the reversible inactivation, maintaining in some cases full enzyme activity at 60 degrees C. A mechanism for the inactivation process, which permitted the reproduction of experimental results, was proposed. The products rhamnose (inhibition constant, 2.1 mM) and prunin (2.6 mM) competitively inhibited the enzyme reaction. The maximum hydrolysis of supersaturated naringin solution, without enzyme inactivation, was observed at 60 degrees C. Hydrolysis of naringin reached 99% with 1% naringin solution, although the hydrolysis degree of naringin was only 40% due to products inhibition when the initial concentration of flavonoid was 10%. The experimental results fitted an equation based on the integrated Michaelis-Menten's, including competitive inhibition by products satisfactorily.
Soria
Fernando
F
Chemical Industry Research Institute (INIQUI), National University of Salta-CONICET, Buenos Aires, Argentina.
Ellenrieder
Guillermo
G
D000465:Algorithms; D001230:Aspergillus / Q000187:drug effects* / Q000201:enzymology; D003432:Cross-Linking Reagents; D003470:Culture Media; D044950:Flavanones; D005419:Flavonoids / Q000378:metabolism*; D005916:Globulins / Q000494:pharmacology; D006026:Glycoside Hydrolases / Q000037:antagonists & inhibitors*; D006358:Hot Temperature; D006868:Hydrolysis; D007700:Kinetics; D010940:Plant Proteins; D013696:Temperature
D003432:Cross-Linking Reagents; D003470:Culture Media; D044950:Flavanones; D005419:Flavonoids; D005916:Globulins; D010940:Plant Proteins; C039828:prunin protein, Prunus; D006026:Glycoside Hydrolases; C022525:alpha-L-rhamnosidase; C005274:naringin