The modular organization of multifunctional peptide synthetases

Joachim Vater, Torsten Stein, Dirk Vollenbroich, Volker Kruft, Brigitte Wittmann-Liebold, Peter Franke, Li Liu, Peter Zuber

Research output: Contribution to journalArticle

Abstract

Gramicidin S synthetase 2 from B. brevis was affinity labeled at its valine thiolation center with the thiol reagent N-[3H]ethylmaleimide. From a tryptic digest of the enzyme-inhibitor complex a radioactive fragment was isolated in pure form by two reversed-phase HPLC steps. It was identified by liquid-phase N-terminal sequencing in combination with electrospray mass spectrometry (ESI-MS) as a hexadecapeptide containing the thiolation motif LGG(H/D)S(L/I). By ESI-MS it was demonstrated that a 4'-phosphopantetheine cofactor was attached to this fragment at its reactive serine. These results are consistent with the 'Multiple Carrier Model' of nonribosomal peptide biosynthesis. Site-specific mutagenesis has been performed in thiolation, elongation, and epimerization motifs of some of the modules of surfactin synthetase from B. subtilis to clarify the function of prominent conserved amino acid residues in the intermediate steps of peptide biosynthesis. The modular structure of multifunctional peptide synthetases is discussed.

Original languageEnglish (US)
Pages (from-to)557-564
Number of pages8
JournalJournal of Protein Chemistry
Volume16
Issue number5
DOIs
StatePublished - 1997
Externally publishedYes

Fingerprint

Nucleic Acid-Independent Peptide Biosynthesis
Peptide Biosynthesis
Peptide Synthases
Sulfhydryl Reagents
Ethylmaleimide
Electrospray Ionization Mass Spectrometry
Biosynthesis
Valine
Enzyme Inhibitors
Site-Directed Mutagenesis
Serine
Peptides
High Pressure Liquid Chromatography
Amino Acids
Mutagenesis
Mass spectrometry
Elongation
Amino acids
Liquids
Enzymes

Keywords

  • Active site mutagenesis
  • Electrospray mass spectrometry
  • Modular structure
  • Multiple 4'-phosphopantetheine cofactors
  • Peptide synthetases
  • Thioester binding site

ASJC Scopus subject areas

  • Biochemistry

Cite this

Vater, J., Stein, T., Vollenbroich, D., Kruft, V., Wittmann-Liebold, B., Franke, P., ... Zuber, P. (1997). The modular organization of multifunctional peptide synthetases. Journal of Protein Chemistry, 16(5), 557-564. https://doi.org/10.1023/A:1026386100259

The modular organization of multifunctional peptide synthetases. / Vater, Joachim; Stein, Torsten; Vollenbroich, Dirk; Kruft, Volker; Wittmann-Liebold, Brigitte; Franke, Peter; Liu, Li; Zuber, Peter.

In: Journal of Protein Chemistry, Vol. 16, No. 5, 1997, p. 557-564.

Research output: Contribution to journalArticle

Vater, J, Stein, T, Vollenbroich, D, Kruft, V, Wittmann-Liebold, B, Franke, P, Liu, L & Zuber, P 1997, 'The modular organization of multifunctional peptide synthetases', Journal of Protein Chemistry, vol. 16, no. 5, pp. 557-564. https://doi.org/10.1023/A:1026386100259
Vater J, Stein T, Vollenbroich D, Kruft V, Wittmann-Liebold B, Franke P et al. The modular organization of multifunctional peptide synthetases. Journal of Protein Chemistry. 1997;16(5):557-564. https://doi.org/10.1023/A:1026386100259
Vater, Joachim ; Stein, Torsten ; Vollenbroich, Dirk ; Kruft, Volker ; Wittmann-Liebold, Brigitte ; Franke, Peter ; Liu, Li ; Zuber, Peter. / The modular organization of multifunctional peptide synthetases. In: Journal of Protein Chemistry. 1997 ; Vol. 16, No. 5. pp. 557-564.
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