TY - JOUR
T1 - Substrate specificity of glutamyl endopeptidase (GE)
T2 - Hydrolysis studies with a bovine α-casein preparation
AU - Kalyankar, Phanindra
AU - Zhu, Yishen
AU - O'Keeffe, Martina
AU - O'Cuinn, Gerard
AU - Fitzgerald, Richard J.
N1 - Copyright © 2012 Elsevier Ltd. All rights reserved.
PY - 2013/1/15
Y1 - 2013/1/15
N2 - Glutamyl endopeptidase (GE) from Alcalase™ 2.4 L was purified using hydrophobic interaction (HIC) and ion-exchange (IEX) chromatography. The yield of GE obtained was approximately 42%. Bovine α-casein (containing αs1- and αs2-casein) was digested with GE at 37 and 50 °C for 4 h. Samples were withdrawn at various time intervals and the peptides generated were analysed using mass spectrometry. GE activity was highly specific and hydrolysed the peptide bond predominantly on the carboxy side of Glu residues while hydrolysis on the carboxyl side of Asp residues was also observed. Hydrolysis did not occur when Pro was at the P1′ position. In Glu-Glu-X (X = Arg, Asn, Ile and Ser) and Glu-Glu-Glu-Lys sequences, hydrolysis of Glu-X and Glu-Lys was preferred. The results are relevant to our understanding of the hydrolytic specificity of Alcalase, a food-grade proteolytic preparation containing GE activity which is used in the generation of casein hydrolysates.
AB - Glutamyl endopeptidase (GE) from Alcalase™ 2.4 L was purified using hydrophobic interaction (HIC) and ion-exchange (IEX) chromatography. The yield of GE obtained was approximately 42%. Bovine α-casein (containing αs1- and αs2-casein) was digested with GE at 37 and 50 °C for 4 h. Samples were withdrawn at various time intervals and the peptides generated were analysed using mass spectrometry. GE activity was highly specific and hydrolysed the peptide bond predominantly on the carboxy side of Glu residues while hydrolysis on the carboxyl side of Asp residues was also observed. Hydrolysis did not occur when Pro was at the P1′ position. In Glu-Glu-X (X = Arg, Asn, Ile and Ser) and Glu-Glu-Glu-Lys sequences, hydrolysis of Glu-X and Glu-Lys was preferred. The results are relevant to our understanding of the hydrolytic specificity of Alcalase, a food-grade proteolytic preparation containing GE activity which is used in the generation of casein hydrolysates.
KW - Bovine α-casein
KW - Glutamyl endopeptidase
KW - LC-MS
KW - Substrate specificity
UR - http://www.scopus.com/inward/record.url?scp=84869077096&partnerID=8YFLogxK
U2 - 10.1016/j.foodchem.2012.08.038
DO - 10.1016/j.foodchem.2012.08.038
M3 - Article
C2 - 23122090
AN - SCOPUS:84869077096
SN - 0308-8146
VL - 136
SP - 501
EP - 512
JO - Food Chemistry
JF - Food Chemistry
IS - 2
ER -