Penicillin amidase

About: Penicillin amidase is a research topic. Over the lifetime, 576 publications have been published within this topic receiving 15563 citations. The topic is also known as: penicillin amidohydrolase & ampicillin acylase.

A novel support of MCM-48 molecular sieve for immobilization of penicillin G acylase

Abstract: As a novel support of immobilizing penicillin G acylase (PGA), MCM-48 and Co-MCM-48 molecular sieves were synthesized and characterized by XRD, N2 adsorption, NH3-TPD, FT-IR and so on. The studies show that MCM-48 and Co-MCM-48 has well ordered long-range structure, narrow pore size distribution, larger surface area and higher concentration of the weakly acidic silanol groups on their surface. Penicillin G acylase was immobilized on MCM-48 or Co-MCM-48 by interacting silanol groups on the surface. The presence of cobalt in the framework of MCM-48 increases the amount of the weak acid sites. For the hydrolysis of penicillin G catalyzed by PGA/Co-MCM-48 (Co/Si=0.01), its specific activity reaches 1682 U/g. After used for six cycles, PGA/MCM-48(0.01) can keep 1375 U/g of the specific activity. If MCM-41 was used as the support, the activity of immobilized PGA is only 402 U/g.

Combinatorial formulation of biocatalyst preparations for increased activity in organic solvents: Salt activation of penicillin amidase

Abstract: A combinatorial experimental technique was used to identify salts and salt mixtures capable of activating penicillin amidase in organic solvents for the transesterification of phenoxyacetate methyl ester with 1-propanol. Penicillin amidase was lyophilized in the presence of various chloride and acetate salts within 96-deep-well plates and catalytic rates measured to determine lead candidates for highly salt-activated preparations. The kinetics of the most active formulations were then further evaluated. These studies revealed that a formulation consisting of 98% (w/w) of a 1:1 KAc:CsCl salt mixture, 1% (w/w) enzyme, and 1% (w/w) potassium phosphate buffer was approximately 35,000-fold more active than the salt-free formulation in hexane, as reflected in values of V(max)/K(m). This extraordinary activation could be extended to more polar solvents, including tert-amyl alcohol, and to formulations with lower total salt contents. A correlation was found between the kosmotropic/chaotropic behavior of the salts (as measured by the Jones-Dole B coefficients) and the observed activation. Strongly chaotropic cations combined with strongly kosmotropic anions yielded the greatest activation, and this is likely due to the influence of the ions on protein-water and protein-salt interactions.

Effects of site-directed mutations on processing and activities of penicillin G acylase from Escherichia coli ATCC 11105.

Abstract: Penicillin G acylase from Escherichia coli ATCC 11105 is synthesized from its precursor polypeptide into a catalytically active heterodimer via a complex posttranslational processing pathway. Substitutions in the pair of aminoacyl residues at the cleavage site for processing the small and large subunits were made. Their processing phenotypes and penicillin G acylase activities were analyzed. By the introduction of a prolyl residue at either position, the processing of the small subunit was blocked without a change in enzymatic activity. Four other substitutions had no effect. At the site for processing the large subunit, four substitutions out of the seven examined blocked processing. In general, penicillin G acylase activity seemed to be proportional to the efficiency of the large-subunit-processing step. Ser-290 is an amino acid critical for processing and also for the enzymatic activity of penicillin G acylase. In the mutant pAATC, in which Ser-290 is mutated to Cys, the precursor is processed, but there is no detectable enzymatic activity. This suggests that there is a difference in the structural requirements for the processing pathway and for enzymatic activity. Recombination analysis of several mutants demonstrated that the small subunit can be processed only when the large subunit is processed first. Some site-directed mutants from which signal peptides were removed showed partial processing phenotypes and reduced enzymatic activities. Their expression showed that the prerequisite for penicillin G acylase activity is the efficient processing of the large subunit and that the maturation of the small subunit does not affect the enzymatic activity.