Assuntos
Penicilina Amidase/química , Bacillus/enzimologia , Sítios de Ligação , Cristalização , Cristalografia por Raios X , Escherichia coli/enzimologia , Evolução Molecular , Ligação de Hidrogênio , Hidrolases/química , Modelos Moleculares , Conformação Proteica , Estrutura Secundária de Proteína , Relação Estrutura-AtividadeRESUMO
Penicillin V acylase from Bacillus sphaericus was purified to homogeneity with an overall yield of 15%. The enzyme exhibited comparatively high specificity for penicillin V, penicillin G, and other related compounds being hydrolyzed at less than 10% of the rate of penicillin V. Moreover, the high rate of hydrolysis was observed when the side chain of the substrate molecule was unsubstituted. Lysine-modifying reagents inactivated the enzyme rapidly. Kinetics and titration studies indicated the involvement of lysine in the catalytic activity of the enzyme.
Assuntos
Bacillus/enzimologia , Penicilina Amidase/isolamento & purificação , Sítios de Ligação , Penicilina Amidase/química , Penicilina Amidase/metabolismo , Especificidade por SubstratoRESUMO
Escherichia coli cells with penicillin acylase activity were sequentially treated at pH 7.8 with aqueous solutions of N-cetyl-N,N,N-trimethylammonium bromide and glutaraldehyde and then immobilized within porous polyacrylamide beads. The immobilized whole cells showed enhanced hydrolysis rates in the conversion of benzylpenicillin to 6-aminopenicillanic acid (6-APA) compared to untreated cells immobilized and used under identical conditions. The immobilized system showed no apparent loss in enzyme activity when used repeatedly over 90 cycles for 6-APA production from 4% benzylpenicillin.
