Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 1994 Jul 15;301(Pt 2):367–372. doi: 10.1042/bj3010367

Substrate specificity of L-delta-(alpha-aminoadipoyl)-L-cysteinyl-D-valine synthetase from Cephalosporium acremonium: demonstration of the structure of several unnatural tripeptide products.

J E Baldwin 1, C Y Shiau 1, M F Byford 1, C J Schofield 1
PMCID: PMC1137089  PMID: 8042979

Abstract

Potential substrates for L-delta-(alpha-aminoadipoyl)-L-(cysteinyl)-D-valine (ACV) synthetase were initially identified using both the amino-acid-dependent ATP<-->pyrophosphate exchange reaction catalysed by the enzyme and the incorporation of 14C-radiolabelled cysteine and valine into potential peptide products. S-Carboxymethylcysteine was an effective substitute for alpha-aminoadipate and both allylglycine and vinylglycine could substitute for cysteine, indicating that the thiol group of cysteine is not essential for peptide formation. L-allo-Isoleucine but not L-isoleucine substituted effectively for valine. The structures of the presumed peptide products derived from these amino acids were confirmed by combined use of electrospray-ionization m.s. (e.s.m.s.) and 1H n.m.r. These results clearly indicate that, in common with other peptide synthetases, but in contrast with ribosomal peptide synthesis, ACV synthetase has a relatively broad substrate specificity.

Full text

PDF

Selected References

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

  1. Baldwin J. E., Bird J. W., Field R. A., O'Callaghan N. M., Schofield C. J. Isolation and partial characterisation of ACV synthetase from Cephalosporium acremonium and Streptomyces clavuligerus. J Antibiot (Tokyo) 1990 Aug;43(8):1055–1057. doi: 10.7164/antibiotics.43.1055. [DOI] [PubMed] [Google Scholar]
  2. Baldwin J. E., Bird J. W., Field R. A., O'Callaghan N. M., Schofield C. J., Willis A. C. Isolation and partial characterisation of ACV synthetase from Cephalosporium acremonium and Streptomyces clavuligerus. Evidence for the presence of phosphopantothenate in ACV synthetase. J Antibiot (Tokyo) 1991 Feb;44(2):241–248. doi: 10.7164/antibiotics.44.241. [DOI] [PubMed] [Google Scholar]
  3. Baldwin J. E., Haber S. B., Hoskins C., Kruse L. I. Synthesis of beta,gamma-unsaturated amino acids. J Org Chem. 1977 Apr 1;42(7):1239–1241. doi: 10.1021/jo00427a031. [DOI] [PubMed] [Google Scholar]
  4. Huffman G. W., Gesellchen P. D., Turner J. R., Rothenberger R. B., Osborne H. E., Miller F. D., Chapman J. L., Queener S. W. Substrate specificity of isopenicillin N synthase. J Med Chem. 1992 May 15;35(10):1897–1914. doi: 10.1021/jm00088a028. [DOI] [PubMed] [Google Scholar]
  5. Igloi G. L., von der Haar F., Cramer F. Aminoacyl-tRNA synthetases from yeast: generality of chemical proofreading in the prevention of misaminoacylation of tRNA. Biochemistry. 1978 Aug 22;17(17):3459–3468. doi: 10.1021/bi00610a006. [DOI] [PubMed] [Google Scholar]
  6. Jensen S. E., Wong A., Rollins M. J., Westlake D. W. Purification and partial characterization of delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase from Streptomyces clavuligerus. J Bacteriol. 1990 Dec;172(12):7269–7271. doi: 10.1128/jb.172.12.7269-7271.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Kleinkauf H., von Döhren H. Nucleic acid independent synthesis of peptides. Curr Top Microbiol Immunol. 1981;91:129–177. doi: 10.1007/978-3-642-68058-8_6. [DOI] [PubMed] [Google Scholar]
  8. Schwecke T., Aharonowitz Y., Palissa H., von Döhren H., Kleinkauf H., van Liempt H. Enzymatic characterisation of the multifunctional enzyme delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase from Streptomyces clavuligerus. Eur J Biochem. 1992 Apr 15;205(2):687–694. doi: 10.1111/j.1432-1033.1992.tb16830.x. [DOI] [PubMed] [Google Scholar]
  9. Shah A. J., Adlard M. W. Determination of beta-lactams and their biosynthetic intermediates in fermentation media by pre-column derivatisation followed by fluorescence detection. J Chromatogr. 1988 Feb 26;424(2):325–336. doi: 10.1016/s0378-4347(00)81109-3. [DOI] [PubMed] [Google Scholar]
  10. Troonen H., Roelants P., Boon B. RIT 2214, a new biosynthetic penicillin produced by a mutant of Cephalosporium acremonium. J Antibiot (Tokyo) 1976 Dec;29(12):1258–1267. doi: 10.7164/antibiotics.29.1258. [DOI] [PubMed] [Google Scholar]
  11. White R. L., DeMarco A. C., Shapiro S., Vining L. C., Wolfe S. Measurement of delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase activity in Streptomyces clavuligerus by high-performance liquid chromatography after precolumn derivatization with o-phthaldialdehyde. Anal Biochem. 1989 May 1;178(2):399–403. doi: 10.1016/0003-2697(89)90660-x. [DOI] [PubMed] [Google Scholar]
  12. Zhang J. Y., Demain A. L. Purification from Cephalosporium acremonium of the initial enzyme unique to the biosynthesis of penicillins and cephalosporins. Biochem Biophys Res Commun. 1990 Jun 29;169(3):1145–1152. doi: 10.1016/0006-291x(90)92015-r. [DOI] [PubMed] [Google Scholar]
  13. Zhang J., Wolfe S., Demain A. L. Biochemical studies on the activity of delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase from Streptomyces clavuligerus. Biochem J. 1992 May 1;283(Pt 3):691–698. doi: 10.1042/bj2830691. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. van Liempt H., von Döhren H., Kleinkauf H. delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase from Aspergillus nidulans. The first enzyme in penicillin biosynthesis is a multifunctional peptide synthetase. J Biol Chem. 1989 Mar 5;264(7):3680–3684. [PubMed] [Google Scholar]

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

RESOURCES