An Unusual Family of Glycosylated Peptides Isolated from Dendroaspis angusticeps Venom and Characterized by Combination of Collision Induced and Electron Transfer Dissociation

Abstract

This study describes the structural characterization of a totally new family of peptides from the venom of the snake green mamba (Dendroaspis angusticeps). Interestingly, these peptides differ in several points from other already known mamba toxins. First of all, they exhibit very small molecular masses, ranging from 1.3 to 2.4 kDa. The molecular mass of classical mamba toxins is in the range of 7 to 25 kDa. Second, the new peptides do not contain disulfide bonds, a posttranslational modification commonly encountered in animal toxins. The third difference is the very high proportion of proline residues in the sequence accounting for about one-third of the sequence. Finally, these new peptides reveal a carbohydrate moiety, indicating a glycosylation in the sequence. The last two features have made the structural characterization of the new peptides by mass spectrometry a real analytical challenge. Peptides were characterized by a combined use of MALDI-TOF/TOF and nanoESI-IT-ETD experiments to determine not only the peptide sequence but also the composition and the position of the carbohydrate moiety. Anyway, such small glycosylated and proline-rich toxins are totally different from any other known snake peptide and form, as a consequence, a new family of peptides.

Venom Down Under: Dynamic Evolution of Australian Elapid Snake Toxins

“…Recently, a proline-rich peptide was isolated from the venom of Dendroaspis angusticeps [54] which was described as being of an unknown peptide or protein type. The results of this study reveal that this proline-rich peptide is part of the propeptide region of the natriuretic peptide precursor, a fact missed by the aforementioned Dendroaspis study despite an Oxyuranus precursor being available that showed significant similarity to this peptide in an early stretch of the propeptide region.…”

Glycopeptide Analysis, Recent Developments and Applications

“…Characterization of Endogenous Glycoproteins-One interesting example of the need for characterization of biologically relevant, isolated glycoproteins is the analysis of several glycopeptides from the venom of Dendroaspis angusticeps (1). These highly active polypeptide compounds were found to be both glycosylated and rich in proline; both features are unusual in snake venom compounds.…”

“…These highly active polypeptide compounds were found to be both glycosylated and rich in proline; both features are unusual in snake venom compounds. Because the analytes of interest were generally under 4 kDa, no enzymatic digestion was required prior to analysis; and a top-down approach was used for assigning both the protein sequence, the glycosylation site occupancy, and the glycans themselves (1). The species contained small O-linked glycans.…”

“…The species contained small O-linked glycans. In this research, a variety of MS techniques were used, including MALDI and ESI for ionization and CID and ETD for fragmentation (1). This study represents the first step necessary for understanding the structure/function connection for this particular snake's venom, but it also serves as an illustrative example of the need for classifying glycosylation, as a first step, for achieving a better understanding of a glycoprotein of interest.…”

“…Frequently, a digestion strategy must be designed that takes into account the nature of the specific protein to be characterized. In the examples above, no digestion was necessary in one case (1), and in another case, chymotrypsin was shown to be the enzyme of choice (6). Proteinase K was also chosen in one of the cited references (4).…”