TY - JOUR
T1 - Crosslinked penicillin acylase aggregates for synthesis of β-lactam antibiotics in organic medium
AU - Illanes, Andrés
AU - Wilson, Lorena
AU - Caballero, Eduardo
AU - Fernández-Lafuente, Roberto
AU - Guisán, José Manuel
PY - 2006/6
Y1 - 2006/6
N2 - 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.
AB - 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.
KW - Ampicillin
KW - Crosslinked enzyme aggregates
KW - Enzyme immobilization
KW - Organic cosolvents
KW - Penicillin acylase
KW - β-lactam antibiotics
UR - http://www.scopus.com/inward/record.url?scp=33745028153&partnerID=8YFLogxK
U2 - 10.1385/ABAB:133:3:189
DO - 10.1385/ABAB:133:3:189
M3 - Article
C2 - 16720900
AN - SCOPUS:33745028153
VL - 133
SP - 189
EP - 202
JO - Applied Biochemistry and Biotechnology
JF - Applied Biochemistry and Biotechnology
SN - 0273-2289
IS - 3
ER -