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Review
. 2018 Sep 15;3(3):159-162.
doi: 10.1016/j.synbio.2018.09.002. eCollection 2018 Sep.

Substrate specificity of the cypemycin decarboxylase CypD

Wei Ding  1 , Tianlu Mo  1 , Dhanaraju Mandalapu  1 , Qi Zhang  1
Affiliations
Review

Substrate specificity of the cypemycin decarboxylase CypD

Wei Ding et al. Synth Syst Biotechnol. .

Erratum in

  • Erratum regarding previously published articles.
    [No authors listed] [No authors listed] Synth Syst Biotechnol. 2020 Oct 12;5(4):328. doi: 10.1016/j.synbio.2020.10.003. eCollection 2020 Dec. Synth Syst Biotechnol. 2020. PMID: 33102826 Free PMC article.

Abstract

The linaridin antibiotic cypemycin is a ribosomal synthesized and post-translationally modified peptide (RiPP) that possesses potent activity against mouse leukemia cells. This peptide natural product contains an S-[(Z)-2-aminovinyl]-d-cysteine (AviCys) moiety in the C-terminus. Formation of AviCys moiety requires an oxidative decarboxylation of the C-terminal Cys of the precursor peptide CypA, and this process is catalyzed by a flavin-containing protein CypD. In this work, we tested CypD substrate specificity with a series of synthetic oligopeptides. We show that most of the N-terminal sequence of CypA is not required for CypD activity, and the C-terminal three residues serve as the minimal structural element for enzyme recognition. We also show that CypD tolerates various substrates with modified C-termini, allowing for the generation of four novel cypemycin variants with modified AviCys moiety by site direct mutagenesis of the precursor peptide CypA. Our study demonstrates the relaxed substrate specificity of CypD and lays a foundation for future bioengineering of AviCys-containing natural products.

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Figures

Fig. 1
Fig. 1
Biosynthesis of cypemycin. (A) Structure of cypemycin. The amino acid sequence of the precursor peptide CypA is shown. Negative numbers represent the position of amino acids within the leader peptide with respect to the first amino acid in the core region (highlighted in yellow). Dehydrobutyrine and AviCys moiety are shown in blue and red. Cypemycin also contains two alloisoleucine (a-Ile) residues that are shown in green. PTM represents posttranslational modifications. (B) The CypD-catalyzed oxidative decarboxylation of the C-terminal Cys, a key step in the formation of the cypemycin AviCys moiety. The blue line represents the peptide chain, and X represents either the CypA Cys19 or a dehydroalanine residue generated by dithiolation of Cys19.
Fig. 2
Fig. 2
LC-HR-MS characterization of (A) V21A, (B) V21C, (C) V21I, and (D) L21I variants of cypemycin. For the detailed HR-MS/MS spectra, see Figs. S11–S14.
See this image and copyright information in PMC

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