Dataset Information

Cleavage of Peptides from Amphibian Skin Revealed by Combining Analysis of Gland Secretion and in Situ MALDI Imaging Mass Spectrometry.

ABSTRACT: Peptides from skin secretions of amphibians are considered important components of their immune system and also play a relevant role in their defense mechanism against predators. Herein, by using mass spectrometry (MS), we characterize the sequence of 13 peptides from the gland secretion of the hylid tree frog, Boana punctata. Using in situ matrix-assisted laser desorption ionization imaging MS of a transverse section of the skin tissue, we show that some peptides are stored as longer molecules that are cleaved after being secreted, whereas others do not undergo any modification. Sequence comparison with peptides from other Boana species and analysis of the three-dimensional theoretical structure indicate that this cleavage depends on both the presence of a specific sequence motif and the secondary structure. The fact that peptides undergo a rapid cleavage upon secretion suggests that stored and secreted peptides may have distinct roles for anuran survival, including defense against pathogens and predators.

SUBMITTER: Brunetti AE

PROVIDER: S-EPMC6044630 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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Publications

Cleavage of Peptides from Amphibian Skin Revealed by Combining Analysis of Gland Secretion and in Situ MALDI Imaging Mass Spectrometry.

Brunetti Andrés E AE Marani Mariela M MM Soldi Rafael A RA Mendonça Jacqueline Nakau JN Faivovich Julián J Cabrera Gabriela M GM Lopes Norberto P NP

ACS omega 20180521 5

Peptides from skin secretions of amphibians are considered important components of their immune system and also play a relevant role in their defense mechanism against predators. Herein, by using mass spectrometry (MS), we characterize the sequence of 13 peptides from the gland secretion of the hylid tree frog, <i>Boana punctata</i>. Using in situ matrix-assisted laser desorption ionization imaging MS of a transverse section of the skin tissue, we show that some peptides are stored as longer mol ...[more]

PMID: 30023919

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Project description:A method of analysis is presented that utilizes matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) to monitor the kinetics and products of RNA cleavage, by use of a program designed to mass-match observed MS peaks with predicted RNA cleavage products. The method is illustrated through application to the study of targeted oxidation of RNA stem loops from HIV-1 Rev Response Element mRNA (RRE RNA) and ribosomal 16S A-site RNA (16S RNA) by metallonucleases. Following incubation of each RNA with catalysts and/or redox co-reactants, reaction mixtures were desalted, and MALDI-TOF MS was used to monitor both time-resolved formation of cleavage products and disappearance of full-length RNA. For each RNA, a unique list was generated that contained the predicted masses of both the full-length, and all of the possible RNA cleavage fragments that resulted from the combination of all possible cleavage sites and each of the six expected overhangs formed at nascent termini adjacent to the cleavage sites. The overhangs corresponded to 2',3'-cyclic phosphate, 3'-phosphate, 3'-phosphoglycolate, 5'- hydroxyl and 5'- phosphate, which corresponded to differing oxidative, hydrolytic, and/or 2'-OH-mediated-endonucleolytic modes of scission. Each mass spectrum was compared with a corresponding list of predicted masses, and peaks were rapidly assigned by use of a Perl script, with a mass-matching tolerance of 200 ppm. Both time-dependent cleavage mediated by metallonucleases and MALDI-TOF-induced fragmentation were observed, and these were distinguished by time-dependent experiments. The resulting data allowed a semi-quantitative assessment of the rate of formation of each overhang at each nucleotide position. Limitations included artifactual skewing of quantification by mass bias, a limited mass range for quantification, and a lack of detection of secondary cleavage products. Nevertheless, the method presented herein provides a rapid, accurate, highly-detailed and semi-quantitative analysis of RNA cleavage that should be widely applicable.

Dataset Information

Cleavage of Peptides from Amphibian Skin Revealed by Combining Analysis of Gland Secretion and in Situ MALDI Imaging Mass Spectrometry.

Publications

Cleavage of Peptides from Amphibian Skin Revealed by Combining Analysis of Gland Secretion and in Situ MALDI Imaging Mass Spectrometry.

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