Lasioglossin LL-I

A novel antimicrobial peptide designated melectin was isolated from the venom of the cleptoparasitic bee Melecta albifrons. Its primary sequence was established as H-Gly-Phe-Leu-Ser-Ile-Leu-Lys-Lys-Val-Leu-Pro-Lys-Val-Met-Ala-His-Met-Lys-NH(2) by Edman degradation and ESI-QTOF mass spectrometry. Synthetic melectin exhibited antimicrobial activity against both gram-positive and -negative bacteria and it degranulated rat peritoneal mast cells, but its hemolytic activity was low. The CD spectra of melectin measured in the presence of trifluoroethanol and sodium dodecyl sulfate showed a high content alpha-helices, which indicates that melectin can adopt an amphipathic alpha-helical secondary structure in an anisotropic environment such as the bacterial cell membrane. To envisage the role of the proline residue located in the middle of the peptide chain on biological activity and secondary structure, we prepared several melectin analogues in which the Pro11 residue was either replaced by other amino acid residues or was omitted. The results of biological testing suggest that a Pro kink in the alpha-helical structure of melectin plays an important role in selectivity for bacterial cells. In addition, a series of N- and C-terminal-shortened analogues was synthesized to examine which region of the peptide is related to antimicrobial activity.

Two novel antimicrobial peptides, named halictines, were isolated from the venom of the eusocial bee Halictus sexcinctus. Their primary sequences were established by ESI-QTOF mass spectrometry, Edman degradation and enzymatic digestion as Gly-Met-Trp-Ser-Lys-Ile-Leu-Gly-His-Leu-Ile-Arg-NH2 (HAL-1), and Gly-Lys-Trp-Met-Ser-Leu-Leu-Lys-His-Ile-Leu-Lys-NH2 (HAL-2). Both peptides exhibited potent antimicrobial activity against Gram-positive and Gram-negative bacteria but also noticeable hemolytic activity. The CD spectra of HAL-1 and HAL-2 measured in the presence of trifluoroethanol or SDS showed ability to form an amphipathic alpha-helical secondary structure in an anisotropic environment such as bacterial cell membrane. NMR spectra of HAL-1 and HAL-2 measured in trifluoroethanol/water confirmed formation of helical conformation in both peptides with a slightly higher helical propensity in HAL-1. Altogether, we prepared 51 of HAL-1 and HAL-2 analogs to study the effect of such structural parameters as cationicity, hydrophobicity, alpha-helicity, amphipathicity, and truncation on antimicrobial and hemolytic activities. The potentially most promising analogs in both series are those with increased net positive charge, in which the suitable amino acid residues were replaced by Lys. This improvement basically relates to the increase of antimicrobial activity against pathogenic Pseudomonas aeruginosa and to the mitigation of hemolytic activity.

The venom from the solitary bee Osmia rufa (Hymenoptera: Megachilidae) was analyzed using mass spectrometry (MS)-based techniques. Sensitive proteomic methods such as on-line LC-ESI-MS and nanoESI-MS analyses revealed more than 50 different compounds with molecular masses ranging from 400 to 4000Da. The major component has a monoisotopic molecular mass of 1924.20Da and its amino acid sequence was elucidated by de novo sequencing using tandem mass spectrometry and Edman degradation. This 17-residue cysteine-free peptide, named osmin, shows some similarities with the mast cell degranulation (MCD) peptide family. Free acid and C-terminally amidated osmins were chemically synthesized and tested for antimicrobial and haemolytic activities. The synthetic C-amidated peptide (native osmin) was found to be about three times more haemolytic than its free acid counterpart, but both peptides are much less lytic than melittin from social bee venom. Preliminary antimicrobial and antifungal tests indicate that both peptides are able to inhibit bacterial and fungal growth at micromolar concentrations.