Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0033036 (APC)
 10,214 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Heterologous production of the heterodimeric penicillin G amidase (PAC) from Providencia rettgeri was optimized in Saccharomyces cerevisiae. Several factors, including the effect of different growth and induction conditions, were identified to be critical for the enzyme overproduction and secretion. The PAC yield was significantly increased by more than 500-fold compared to that obtained in the native bacterium, and the recombinant enzyme was almost entirely secreted. Electrophoretic characterization of the secreted rPAC(Pr), which was purified over 20-fold by a combination of hydrophobic interaction and ion-exchange chromatography, demonstrated a microheterogeneity of the recombinant enzyme. The recombinant PAC(Pr) was further characterized in terms of specific activity, pH, and temperature profiles and kinetic parameters. The data presented here suggest that by overexpressing rPAC(Pr) in S.cerevisiae and purifying secreted enzyme from culture medium one can readily obtain a large amount of an alternative source of penicillin amidase with properties comparable to that of todays main industrial source of enzyme.
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PMID:High-level secretory expression of penicillin amidase from Providencia rettgeri in Saccharomyces cerevisiae: purification and characterization. 1193 4

Penicillin G acylase (PAC; EC 3.5.1.11) is the key enzyme used in the industrial production of beta-lactam antibiotics. This enzyme hydrolyzes the side chain of penicillin G and related beta-lactam antibiotics releasing 6-amino penicillanic acid (6-APA), which is the building block in the manufacture of semisynthetic penicillins. PAC from Escherichia coli strain ATCC 11105, Bacillus megaterium strain ATCC 14945 and mutants of these two strains is currently used in industry. Genes encoding for PAC from various bacterial sources have been cloned and overexpressed with significant improvements in transcription, translation and post-translational processing. Recent developments in enzyme engineering have shown that PAC can be modified to gain conformational stability and desired functionality. This review provides an overview of recent advances in the production, stabilization and application of PAC, highlighting the recent biotechnological approaches for the improved catalysis of PAC.
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PMID:Recent biotechnological interventions for developing improved penicillin G acylases. 1623 81

There is an increasing demand for the development of sensitive enzymatic assays compatible with droplet-based microfluidics. Here we describe an original strategy, activity-fed translation (AFT), based on the coupling of enzymatic activity to in vitro translation of a fluorescent protein. We show that methionine release upon the hydrolysis of phenylacetylmethionine by penicillin acylase enabled in vitro expression of green fluorescent protein. An autocatalytic setup where both proteins are expressed makes the assay highly sensitive, as fluorescence was detected in droplets containing single PAC genes. Adding a PCR step in the droplets prior to the assay increased the sensitivity further. The strategy is potentially applicable for any activity that can be coupled to the production of an amino acid, and as the microdroplet volume is small the use of costly reagents such as in vitro expression mixtures is not limiting for high-throughput screening projects.
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PMID:Activity-Fed Translation (AFT) Assay: A New High-Throughput Screening Strategy for Enzymes in Droplets. 2591 25