Abstract
In the present research effort, production of derivatives of cardosin A (a plant protease) encompassing full stabilization of its dimeric structure has been achieved, via covalent, multi-subunit immobilization onto highly activated agarose-glutaraldehyde supports. Boiling such enzyme derivatives in the presence of sodium dodecyl sulfate and β-mercaptoethanol did not lead to leaching of enzyme, thus providing evidence for the effectiveness of the attachment procedure. Furthermore, the cardosin A derivatives prepared under optimal conditions presented ca. half the specific activity of the enzyme in soluble form, and were successfully employed at laboratory-scale trials to perform (selective) hydrolysis of α-lactalbumin (α-La), one of the major proteins in bovine whey. Hydrolysates of α-La were assayed for by the OPA method, as well as by FPLC, SDS-PAGE and HPLC. Thermal inactivation of the immobilized cardosin A was also assessed at 40, 50 and 55°C; at these temperatures, no thermal denaturation took place during incubation for 48h. The highest degree of hydrolysis was attained by 5h reaction, at 55°C and pH 5.2. SDS-PAGE of α-La hydrolysates displayed bands corresponding to low molecular weight peptides. Our results suggest that cardosin A in immobilized form is a good candidate to bring about proteolysis in the dairy industry, namely in whey processing.
Original language | English |
---|---|
Pages (from-to) | 908-916 |
Number of pages | 9 |
Journal | Enzyme and Microbial Technology |
Volume | 33 |
Issue number | 7 |
DOIs | |
Publication status | Published - 2 Dec 2003 |
Keywords
- Agarose
- Attachment
- Cardosin
- Dairy foods
- Enzyme
- Protease
- Structural stabilization
- α-Lactalbumin