title: Crosslinked penicillin acylase aggregates for synthesis of beta-lactam antibiotics in organic medium.
abstract: Crosslinked enzyme aggregates (CLEAs) of a partially purified penicillin acylase from a recombinant Escherichia coli strain have been produced as a novel type of biocatalysts well endowed to perform in organic media. Different protein precipitants were studied and glutaraldehyde was used as the crosslinking agent. Precipitation curves were obtained for all precipitants to determine the concentrations at which all the protein precipitated out of the solution. The effect of the glutaraldehyde-to-protein ratio was studied with respect to process recovery and the specific activity and stability of the biocatalyst. Recovery of penicillin acylase activity was moderately high, about 50%; major losses of enzyme activity were produced at the precipitation step. Specific activities of all CLEAs were very high, which is one of the advantages of using nonsupported biocatalysts. Ammonium sulfate and tert-butyl alcohol were the best precipitants at a glutaraldehyde-protein mass ratio of 2 and were selected to perform the kinetically controlled synthesis of ampicillin in 60% (v/v) ethylene glycol medium. At comparable conversion yields, volumetric and specific antibiotic productivity were much higher for CLEAs than for carrier-bound penicillin acylases.
authors.affiliation: School of Biochemical Engineering, Pontificia Universidad Católica Valparaíso, PO Box 4059, Valparaíso, Chile. aillanes@ucv.cl
authors.forename: Andrés
authors.initials: A
authors.lastname: Illanes
authors.forename: Lorena
authors.initials: L
authors.lastname: Wilson
authors.forename: Eduardo
authors.initials: E
authors.lastname: Caballero
authors.forename: Roberto
authors.initials: R
authors.lastname: Fernández-Lafuente
authors.forename: José Manuel
authors.initials: JM
authors.lastname: Guisán
mesh_terms: D000900:Anti-Bacterial Agents / Q000138:chemical synthesis*; D002384:Catalysis; D011232:Chemical Precipitation; D003432:Cross-Linking Reagents / Q000737:chemistry; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000737:chemistry; D004926:Escherichia coli / Q000201:enzymology; D019855:Ethylene Glycol / Q000737:chemistry; D007700:Kinetics; D009930:Organic Chemicals / Q000737:chemistry*; D010399:Penicillin Amidase / Q000378:metabolism*; D012997:Solvents / Q000737:chemistry; D047090:beta-Lactams / Q000138:chemical synthesis*; D020002:tert-Butyl Alcohol / Q000737:chemistry
chemical_list: D000900:Anti-Bacterial Agents; D003432:Cross-Linking Reagents; D004800:Enzymes, Immobilized; D009930:Organic Chemicals; D012997:Solvents; D047090:beta-Lactams; D010399:Penicillin Amidase; D019855:Ethylene Glycol; D020002:tert-Butyl Alcohol