Peptidomic analysis of the venom of the solitary bee Xylocopa appendiculata circumvolans

Abstract

BackgroundAmong the hymenopteran insect venoms, those from social wasps and bees – such as honeybee, hornets and paper wasps – have been well documented. Their venoms are composed of a number of peptides and proteins and used for defending their nests and themselves from predators. In contrast, the venoms of solitary wasps and bees have not been the object of further research. In case of solitary bees, only major peptide components in a few venoms have been addressed. Therefore, the aim of the present study was to explore the peptide component profile of the venom from the solitary bee Xylocopa appendiculata circumvolans by peptidomic analysis with using LC-MS.MethodsA reverse-phase HPLC connected to ESI-OrbiTrap MS was used for LC-MS. On-line mass fingerprinting was made from TIC, and data-dependent tandem mass spectrometry gave MSMS spectra. A major peptide component was isolated by reverse-phase HPLC by conventional way, and its sequence was determined by Edman degradation, which was finally corroborated by solid phase synthesis. Using the synthetic specimen, biological activities (antimicrobial activity, mast cell devaluation, hemolysis, leishmanicidal activity) and pore formation in artificial lipid bilayer were evaluated.ResultsOn-line mass fingerprinting revealed that the crude venom contained 124 components. MS/MS analysis gave 75 full sequences of the peptide components. Most of these are related to the major and novel peptide, xylopin. Its sequence, GFVALLKKLPLILKHLH-NH2, has characteristic features of linear cationic α-helical peptides; rich in hydrophobic and basic amino acids with no disulfide bond, and accordingly, it can be predicted to adopt an amphipathic α-helix secondary structure. In biological evaluation, xylopin exhibited broad-spectrum antimicrobial activity, and moderate mast cell degranulation and leishmanicidal activities, but showed virtually no hemolytic activity. Additionally, the peptide was able to incorporate pores in artificial lipid bilayers of azolectin, confirming the mechanism of the cytolytic activity by pore formation in biological membranes.ConclusionsLC-ESI-MS and MS/MS analysis of the crude venom extract from a solitary bee Xylocopa appendiculata circumvolans revealed that the component profile of this venom mostly consisted of small peptides. The major peptide components, xylopin and xylopinin, were purified and characterized in a conventional manner. Their chemical and biological characteristics, belonging to linear cationic α-helical peptides, are similar to the known solitary bee venom peptides, melectin and osmin. Pore formation in artificial lipid bilayers was demonstrated for the first time with a solitary bee peptide.

Functional characterization, antimicrobial effects, and potential antibacterial mechanisms of new mastoparan peptides from hornet venom (Vespa ducalis, Vespa mandarinia, and Vespa affinis)

“…2, we show details of the hydrophobicity and structure of a typical AMP that we identified, U-VVTX-Vm1a from V. ducalis. There are both hydrophilic (N-terminal) and hydrophobic (C-terminal) regions, suggesting that this AMPs could be amphiphilic (Kazuma et al, 2017). Hydrophilic and hydrophobic regions were likewise found in all our AMPs.…”

“…MP peptides extracted from wasp venom can play a beneficial role in clinical medicine. Mastoparan from Polybia paulista can treat parasitic infection (Kazuma et al, 2017). Gupta et al (2021) found that the new AMPs of mastoparan had antibacterial effects on drug-resistant E.coli of newborn calves at a concentration of 2MIC.…”

Venom profile of the European carpenter bee Xylocopa violacea: Evolutionary and applied considerations on its toxin components

“…The few existing studies on solitary bee venom are mostly application-driven and focus on the identification of single compounds in crude venom and their possible bioactivity. A major attention is paid to new antimicrobial peptides from solitary bees ( Čujová et al, 2014 ; Kazuma et al, 2017 ; Ko et al, 2020 ; Monincová et al, 2010 , 2012 , 2014 ; Raghuraman and Chattopadhyay, 2007 ; Stöcklin et al, 2010 ) for which hymenopterans are particular known for ( Moreau, 2013 ; Slaninova et al, 2012 ). The only proteomic study using whole solitary bee venom is the one of the Japanese carpenter bee X. appendiculata circumvolans ( Kazuma et al, 2017 ), however it also focuses on shorter peptides.…”

The Antimicrobial Peptide Melectin Shows Both Antimicrobial and Antitumor Activity via Membrane Interference and DNA Binding

“…At the same time, it can degranulate peritoneal mast cells from rats. 24 However, more information on Melectin is needed before it can be seen as a potent treatment candidate. Does it also affect drug-resistant bacteria?…”