Dereplication and de novo sequencing of nonribosomal peptides - PubMed (original) (raw)

Dereplication and de novo sequencing of nonribosomal peptides

Julio Ng et al. Nat Methods. 2009 Aug.

Abstract

Nonribosomal peptides (NRPs) are of great pharmacological importance, but there is currently no technology for high-throughput NRP 'dereplication' and sequencing. We used multistage mass spectrometry followed by spectral alignment algorithms for sequencing of cyclic NRPs. We also developed an algorithm for comparative NRP dereplication that establishes similarities between newly isolated and previously identified similar but nonidentical NRPs, substantially reducing dereplication efforts.

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Figure 1

Figure 1

Experimental and theoretical spectra of seglitide Cyclic(N-methyl-Ala, Tyr, D-Trp, Lys, Val, Phe (a) Cyclic diagram of seglitide. A+14 denotes a methylated alanine; the integer residue masses are 85, 163, 186, 128, 99 and 147. (b) MS2 fragmentation of seglitide generates up to 6 linear peptides representing different linearized variants of the same cyclic peptide. (c) The theoretical spectrum for seglitide is a superposition of the fragment masses from the linearized peptide. (d) Experimental spectrum of seglitide (the peaks corresponding to prefix masses are shown in red). (e) The auto-convolution of the spectrum in insert d has prominent peaks for offsets corresponding to masses of amino acids (shown in red). The peak at 0 is truncated. (f) Generation of a gapped peptide from a theoretical spectrum of seglitide. The theoretical spectrum is colored to highlight various linear peptides. For illustration purposes only 3 linearized (A+14YWKV (blue), FA+14YWKV (red) and VFA+14YWK (green)) versions of the cyclic peptide are shown. The frequent 2-amino-acid tag YW is observed in 3 different locations in the spectrum. Additionally, the offsets between 3 consecutive locations of tag YW reveal the masses of amino acids F and V. (g) The gapped peptide constructed from f combines YW (derived from a frequent tag) with VF (derived from the inter distances between tag locations). A+14 and K are inferred from the flanking masses of YW and VF. The complete sequence A+14YWKVF is recovered, but gaps may be generated.

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