title: Substrate specificity of glutamyl endopeptidase (GE): hydrolysis studies with a bovine α-casein preparation.
abstract: 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 α(s1)- and α(s2)-casein) was digested with GE at 37 and 50°C for 4h. Samples were withdrawn at various time intervals and the peptides generated were analysed using mass spectrometry.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 P(1)' 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.
authors.affiliation: Department of Life Sciences, University of Limerick, Limerick, Ireland.
authors.forename: Phanindra
authors.initials: P
authors.lastname: Kalyankar
authors.forename: Yishen
authors.initials: Y
authors.lastname: Zhu
authors.forename: Martina
authors.initials: M
authors.lastname: O'Keeffe
authors.forename: Gerard
authors.initials: G
authors.lastname: O'Cuinn
authors.forename: Richard J
authors.initials: RJ
authors.lastname: FitzGerald
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001407:Bacillus / Q000737:chemistry / Q000201:enzymology*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D002364:Caseins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002417:Cattle; D006868:Hydrolysis; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D002364:Caseins; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase