Topic
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.
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TL;DR: In this paper, bis(phenylacet)amides with penicillin amidase from E. coli (EC 3.5.1.11) were synthesized starting from cheap carbocyclic olefins and cis-diepoxy derivatives.
31 citations
31 citations
TL;DR: It is shown that a heterodimeric enzyme can be expressed as a fusion protein that matures to an active biocatalyst connected to the coat protein of phage fd, resulting in a phage to which the beta-sub unit is covalently linked and the alpha-subunit is non-covalently attached.
Abstract: The large heterodimeric penicillin G acylase from Alcaligenes faecalis was displayed on the surface of phage fd. We fused the coding sequence (alpha subunit-internal peptide-beta subunit) to the gene of a phage coat protein. A modified g3p signal sequence was used to direct the polypeptide to the periplasm. Here we show that a heterodimeric enzyme can be expressed as a fusion protein that matures to an active biocatalyst connected to the coat protein of phage fd, resulting in a phage to which the beta-subunit is covalently linked and the alpha-subunit is non-covalently attached. The enzyme can be displayed either fused to the minor coat protein g3p or fused to the major coat protein g8p. In both cases the penicillin G acylase on the phage has the same Michaelis constant as its freely soluble counterpart, indicating a proper folding and catalytic activity of the displayed enzyme. The display of the heterodimer on phage not only allows its further use in protein engineering but also offers the possibility of applying this technology for the excretion of the enzyme into the extracellular medium, facilitating purification of the protein. With the example of penicillin acylase the upper limit for a protein to become functionally displayed by phage fd has been further explored. Polyvalent display was not observed despite the use of genetic constructs designed for this aim. These results are discussed in relation to the pore size being formed by the g4p multimer.
31 citations
TL;DR: In this article, the effects of temperature, polymer molecular weight, and polymer and salt concentrations on the partitioning of penicillin G in aqueous two-phase systems were studied.
Abstract: The partitioning of penicillin G acylase in aqueous two-phase systems (ATPS’s) containing poly(ethylene glycol) (PEG) 20000 or 35000 and potassium dihydrogen phosphate (KH2PO4) or sodium citrate (C6H5Na3O7·5H2O) has been measured at three temperatures, (301.2, 307.2, and 310.2) K. The effects of temperature, polymer molecular weight, and polymer and salt concentrations on the partitioning of penicillin G in the ATPS were studied. The experimental data showed that the composition of salt has a large effect on partitioning of penicillin G in ATPS, and the temperature of the system has a small effect on the partitioning. The UNIFAC-FV group contribution model (Pazuki et al., Ind. Eng. Chem. Res. 2009, 48, 4109−4118) was used in correlating the partition coefficients of penicillin G in polymer + ATPS.
31 citations
TL;DR: A model was developed for predicting the equilibrium conversion of Pen G quite reasonably for different values of pH, initial penicillin G concentration and phase volume ratio and it was found that this conversion can exceed 90% because crystallization of 6-aminopenicillanic acid shifts the equilibrium to the product side.
Abstract: In the present downstream processing of penicillin G, penicillin G is extracted from the fermentation broth with an organic solvent and purified as a potassium salt via a number of back-extraction and crystallization steps. After purification, penicillin G is hydrolyzed to 6-aminopenicillanic acid, a precursor for many semisynthetic beta-lactam antibiotics. We are studying a reduction in the number of pH shifts involved and hence a large reduction in the waste salt production. To this end, the organic penicillin G extract is directly to be added to an aqueous immobilized enzyme suspension reactor and hydrolyzed by extractive catalysis. We found that this conversion can exceed 90% because crystallization of 6-aminopenicillanic acid shifts the equilibrium to the product side. A model was developed for predicting the equilibrium conversion in batch systems containing both a water and a butyl acetate phase, with either potassium or D-p-hydroxyphenylglycine methyl ester as counter-ion of penicillin G. The model incorporates the partitioning equilibrium of the reactants, the enzymatic reaction equilibrium, and the crystallization equilibrium of 6-aminopenicillanic acid. The model predicted the equilibrium conversion of Pen G quite reasonably for different values of pH, initial penicillin G concentration and phase volume ratio. The model can be used as a tool for optimizing the enzymatic hydrolysis.
31 citations