Skip to main content
Biochemical Journal logoLink to Biochemical Journal
. 1984 Sep 15;222(3):789–795. doi: 10.1042/bj2220789

Purification of isopenicillin N synthetase.

C P Pang, B Chakravarti, R M Adlington, H H Ting, R L White, G S Jayatilake, J E Baldwin, E P Abraham
PMCID: PMC1144243  PMID: 6435606

Abstract

Isopenicillin N synthetase was extracted from Cephalosporium acremonium and purified about 200-fold. The product showed one major protein band, coinciding with synthetase activity, when subjected to electrophoresis in polyacrylamide gel. An isopenicillin N synthetase from Penicillium chrysogenum was purified about 70-fold by similar procedures. The two enzymes resemble each other closely in their Mr, in their mobility on electrophoresis in polyacrylamide gel and in their requirement for Fe2+ and ascorbate for maximum activity. Preliminary experiments have shown that a similar isopenicillin N synthetase can be extracted from Streptomyces clavuligerus.

Full text

PDF
792

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Berks M., Redhead K., Abraham E. P. Isolation and properties of an inducible and a constitutive beta-lactamase from Pseudomonas aeruginosa. J Gen Microbiol. 1982 Jan;128(1):155–159. doi: 10.1099/00221287-128-1-155. [DOI] [PubMed] [Google Scholar]
  2. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  3. Brewer S. J., Farthing J. E., Turner M. K. The oxygenation of the 3-methyl group of 7beta-(5-D-aminoadipamido)-3-methylceph-3-em-4-carboxylic acid (desacetoxycephalosporin C) by extracts of Acremonium chrysogenum [proceedings]. Biochem Soc Trans. 1977;5(4):1024–1026. doi: 10.1042/bst0051024. [DOI] [PubMed] [Google Scholar]
  4. Davies R. B., Abraham E. P. Separation, purification and properties of beta-lactamase I and beta-lactamase II from Bacillus cereus 569/H/9. Biochem J. 1974 Oct;143(1):115–127. doi: 10.1042/bj1430115. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Fawcett P. A., Usher J. J., Abraham E. P. Behaviour of tritium-labelled isopenicillin N and 6-aminopenicillanic acid as potential penicillin precursors in an extract of Penicillum chrysogenum. Biochem J. 1975 Dec;151(3):741–746. doi: 10.1042/bj1510741. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Fawcett P. A., Usher J. J., Huddleston J. A., Bleaney R. C., Nisbet J. J., Abraham E. P. Synthesis of delta-(alpha-aminoadipyl)cysteinylvaline and its role in penicillin biosynthesis. Biochem J. 1976 Sep 1;157(3):651–660. doi: 10.1042/bj1570651. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hollander I. J., Shen Y. Q., Heim J., Demain A. L., Wolfe S. A pure enzyme catalyzing penicillin biosynthesis. Science. 1984 May 11;224(4649):610–612. doi: 10.1126/science.6546810. [DOI] [PubMed] [Google Scholar]
  8. Kupka J., Shen Y. Q., Wolfe S., Demain A. L. Studies on the ring-cyclization and ring-expansion enzymes of beta-lactam biosynthesis in Cephalosporium acremonium. Can J Microbiol. 1983 May;29(5):488–496. doi: 10.1139/m83-078. [DOI] [PubMed] [Google Scholar]
  9. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  10. Loder P. B., Abraham E. P. Isolation and nature of intracellular peptides from a cephalosporin C-producing Cephalosporium sp. Biochem J. 1971 Jul;123(3):471–476. doi: 10.1042/bj1230471. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. O'Sullivan J., Abraham E. P. The conversion of cephalosporins to 7 alpha-methoxycephalosporins by cell-free extracts of Streptomyces clavuligerus. Biochem J. 1980 Feb 15;186(2):613–616. doi: 10.1042/bj1860613. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. O'Sullivan J., Aplin R. T., Stevens C. M., Abraham E. P. Biosynthesis of a 7-alpha-methoxycephalosporin. Incorporation of molecular oxygen. Biochem J. 1979 Apr 1;179(1):47–52. doi: 10.1042/bj1790047. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Smith B., Warren S. C., Newton G. G., Abraham E. P. Biosynthesis of penicillin N and cephalosporin C. Antibiotic production and other features of the metabolism of Cephalosporium sp. Biochem J. 1967 Jun;103(3):877–890. doi: 10.1042/bj1030877. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. White R. L., John E. M., Baldwin J. E., Abraham E. P. Stoichiometry of oxygen consumption in the biosynthesis of isopenicillin from a tripeptide. Biochem J. 1982 Jun 1;203(3):791–793. doi: 10.1042/bj2030791. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES