Publication: Localization and characterization of inclusion bodies in recombinant Escherichia coli cells overproducing penicillin G acylase
Issued Date
1997-06-04
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ISSN
01757598
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2-s2.0-0030916974
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Mahidol University
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SCOPUS
Bibliographic Citation
Applied Microbiology and Biotechnology. Vol.47, No.4 (1997), 373-378
Suggested Citation
N. Sriubolmas, W. Panbangred, S. Sriurairatana, V. Meevootisom Localization and characterization of inclusion bodies in recombinant Escherichia coli cells overproducing penicillin G acylase. Applied Microbiology and Biotechnology. Vol.47, No.4 (1997), 373-378. doi:10.1007/s002530050943 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/17892
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Title
Localization and characterization of inclusion bodies in recombinant Escherichia coli cells overproducing penicillin G acylase
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Abstract
Various concentrations of isopropyl β-D-thiogalactopyranoside (IPTG) were used to induce production of the enzyme penicillin G acylase by recombinant Escherichia coli harboring plasmid pQEA11. The plasmid pQEA11 carries a wild-type pga gene, which is under the control of the tac promoter and lacI(q). At low IPTG concentrations (0.025-0.1 mM), enzyme activity increased with increasing IPTG concentrations. At higher IPTG concentrations (0.2 and 0.5 mM), enzyme activity declined progressively. Examination of induced recombinant E. coli cells by transmission electron microscopy showed the presence of only periplasmic inclusion bodies at low IPTG concentrations (up to 0.1 mM) and both periplasmic and cytoplasmic inclusion bodies at high IPTG concentrations (0.2 mM and 0.5 mM). Results from sodium dodecyl sulfate/polyacrylamide gel electrophoresis and immunoblots of whole-cell proteins, membrane proteins and inclusion body proteins in these cells indicated that cytoplasmic inclusion bodies constituted an accumulation of pre-proenzyme (i.e., precursor polypeptide containing a signal peptide) and that periplasmic inclusion bodies constituted an accumulation of proenzyme (i.e., precursor polypeptide lacking a signal peptide).