title: Immobilization of angiotensin-converting enzyme on glyoxyl-agarose.
abstract: The assay of angiotensin-converting enzyme (ACE) inhibition by food-derived peptides is usually carried out by using soluble ACE in a batch process. The purification of this enzyme from tissues is not an easy task, and the resulting preparation loses activity very fast. In addition, ACE commercial preparations are very expensive. In this work the immobilization of ACE, through lysine amino groups, to 4% beads cross-linked (4 BCL) glyoxyl-agarose is described. The amount of immobilized enzyme increased with increasing concentrations of enzyme and with incubation time until a saturation point was reached at 50 mg protein/mL gel and 3.5 hours, respectively. The IC50 values for a noncompetitive sunflower peptide inhibitor were similar for the soluble (30.56 microM) and immobilized (32.7 microM) enzymes. An immobilized derivative was obtained that was 60 times more stable than the soluble enzyme at 60 degrees C. This procedure yields a derivative that can be reused and has increased thermal stability compared to that of the soluble enzyme. Thus, ACE immobilization is a good alternative to using soluble freshly prepared or commercial preparations because of economical and practical reasons.
authors.affiliation: Instituto de la Grasa (C.S.I.C.), Padre García Tejero 4, 41012-Sevilla, Spain.
authors.forename: Cristina
authors.initials: C
authors.lastname: Megías
authors.forename: Justo
authors.initials: J
authors.lastname: Pedroche
authors.forename: María
authors.initials: M
authors.lastname: del Mar Yust
authors.forename: Manuel
authors.initials: M
authors.lastname: Alaiz
authors.forename: Julio
authors.initials: J
authors.lastname: Girón-Calle
authors.forename: Francisco
authors.initials: F
authors.lastname: Millán
authors.forename: Javier
authors.initials: J
authors.lastname: Vioque
mesh_terms: D000806:Angiotensin-Converting Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000378:metabolism; D006038:Glyoxylates; D006368:Helianthus / Q000737:chemistry; D006358:Hot Temperature; D010455:Peptides / Q000494:pharmacology; D007703:Peptidyl-Dipeptidase A / Q000378:metabolism; D012685:Sepharose; D012995:Solubility
chemical_list: D000806:Angiotensin-Converting Enzyme Inhibitors; D004800:Enzymes, Immobilized; D006038:Glyoxylates; D010455:Peptides; C070399:glyoxyl agarose; D012685:Sepharose; D007703:Peptidyl-Dipeptidase A