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Plasma desorption mass spectrometry of two synthetic sarafotoxins: Side reactions and characterization of the intermediates
Author(s) -
Lamthanh Hung,
Deprun Claude,
Beyec Yvon Le
Publication year - 1994
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/rcm.1290081213
Subject(s) - chemistry , side chain , methionine , amino acid , mass spectrometry , methionine sulfoxide , peptide , tryptophan , solid phase synthesis , stereochemistry , peptide synthesis , organic chemistry , biochemistry , chromatography , polymer
Sarafotoxins (SRTXs) form a family of toxic and potent vasoconstrictor peptides of 21 amino acids and two disulfide bonds. They are present in the venom of the burrowing asp Atractaspis engaddensis . We have made two derivatives of the amino acid sequence of SRTX‐b, one of the most potent isotoxins, in the solid phase. First, we replaced Ser 2 by Thr, to investigate whether, as previously postulated, this change is responsible for the weak activities of SRTXs c and d. Secondly, we replaced Ser 2 , Asp 18 and Val 19 respectively by Thr, Gly and He, with a view to generating SRTX‐e whose amino acid sequence was deduced from cDNA. Solid‐phase peptide synthesis (SPPS) was performed according to the tert ‐butyloxycarbonyl strategy and the disulfides were paired sequentially using a selective chemistry. The disulfide 1–15 was formed by oxidation of cysteines 1,15 with ferricyanide, whereas disulfide 3–11 was made by iodine oxidation of Acm‐blocked cysteines 3,11 . By plasma desorption mass spectrometry (PDMS), we monitored all possible side reactions that occurred during the synthesis. We thus observed a benzyl shift in mass spectra when aspartic and glutamic acid side chains were protected by a benzyl group during the SPPS. This could be circumvented by using instead, a cyclohexyl protecting group. We also noted the oxidation of the methionine and the tryptophan side chain (formation of methionine sulfoxide and oxindole ring of tryptophan) to a small extent during the cleavage peptide/solid phase oxidation of the methionine side chain during the formation of the disulfide 1–15 by ferricyanide. These known side reactions could be avoided by using a mixture of scavengers (ethanedithiol, dimethylsulfide, thioanisole, m ‐cresol and skatole) and by shortening the ferricyanide oxidation time to one hour. The sequential disulfied bond formation starting from the linear peptide (mono‐disulfied and bi‐disulfide) was also monitored by PDMS. PDMS therefore proved to be a sensitive analytical method for the measurement of the molecular weights of synthetic or natural peptides (SRTX peptides in this application) in the field of peptide and protein chemistry.