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Isolation and sequence analysis of peptides from the skin secretion of the Middle East tree frog Hyla savignyi

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Abstract

Novel peptides were identified in the skin secretion of the tree frog Hyla savignyi. Skin secretions were collected by mild electrical stimulation. Peptides were separated by reversed-phase high-performance liquid chromatography. Mass spectra were acquired by electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS), and fragment ion spectra were obtained after collision-induced dissociation and electron capture dissociation. Peptides were analyzed by manual de novo sequencing and composition-based sequencing (CBS). Sequence analyses of three so far undescribed, structurally unrelated peptides are presented in this paper, having the sequences DDSEEEEVE-OH, P*EEVEEERJK-OH, and GJJDPJTGJVGGJJ-NH2. The glutamate-rich sequences are assumed to be acidic spacer peptides of the prepropeptide. One of these peptides contains the modified amino acid hydroxyproline, as identified and localized by high-accuracy FTICR-MS. Combination of CBS and of experience-based manual sequence analysis as complementary and database-independent sequencing strategies resulted in peptide identification with high reliability.

So-far unknown natural frog skin peptides were identified by high-resolution CID and ECD MS/MS and by composition-based de novo sequencing. Sequences were confirmed by comparison of MS/MS spectra with synthesized analogs

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Acknowledgments

We would like to thank Faraham Ahmadzadeh and Hossein Hashempour from Shahid Beheshti University, Tehran, for collecting the skin secretions. Financial support by the Deutsche Forschungsgemeinschaft (DFG, Sp314/10-1) and by the Bundesministerium für Bildung und Forschung (BMBF, NGFN project 0313442) is gratefully acknowledged.

This publication represents a component of the doctoral (Dr. rer. nat.) thesis of Markus Langsdorf in the Faculty of Biology and Chemistry at the Justus Liebig University Giessen, Germany.

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Authors and Affiliations

  1. Institute of Inorganic and Analytical Chemistry, Justus Liebig University, 35392, Schubertstr. 60, Bldg. 16, Giessen, Germany

    Markus Langsdorf, Andreas Römpp & Bernhard Spengler

  2. Medicinal Plants and Drugs Research Institute, Shahid Beheshti University, Evin, Tehran, 1983963113, Iran

    Alireza Ghassempour

Authors
  1. Markus Langsdorf
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  2. Alireza Ghassempour
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  3. Andreas Römpp
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  4. Bernhard Spengler
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Correspondence to Bernhard Spengler.

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Published in the special issue Mass Spectrometry (DGMS 2010) with Guest Editors Andrea Sinz and Jürgen Schmidt.

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Langsdorf, M., Ghassempour, A., Römpp, A. et al. Isolation and sequence analysis of peptides from the skin secretion of the Middle East tree frog Hyla savignyi . Anal Bioanal Chem 398, 2853–2865 (2010). https://doi.org/10.1007/s00216-010-4131-8

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