PEGylation of Melittin: Structural Characterization and Hemostatic Effects

Abstract

To promote and understand the structure-property relationship for hemostasis, we modified melittin (MLT) using a four-arm poly(ethylene glycol) (PEG) with N-hydroxysuccinimide ester. The PEGylation was characterized by FTIR, MALDI-MS, NMR, a bicinchoninic acid assay, circular dichroism, hemolysis assay, and thromboelastography. Changes in the reaction conditions affected the extent of the modification, the numbers of MLT conjugated to PEG arms, and possible PEGylation sites. The reaction at pH 9.2 with a high MLT/PEG ratio, resulted in the highest modification. Reactions in dimethylsulfoxide (DMSO) resulted in more multi-arm coupled MLT, reaching a maximum of four MLT per PEG. The helicity of the modified peptide, relative to the native peptide, was essentially maintained in DMSO, but substantially lost at pH 9.2. PEGylation reduced the hemolytic effects of MLT and subsequently changed its coagulation profiles. The overall hemostatic effects of MLT modified in DMSO indicate that this may be a convenient approach to the PEGylation of biomolecules for biomedical applications.

Potentiality of Melittin-Loaded Niosomal Vesicles Against Vancomycin-Intermediate Staphylococcus aureus and Staphylococcal Skin Infection

“…Mel showed characteristic absorption peaks at 1657 and 1539 cm −1 , corresponding to the stretching of the C=O and N-H bonds of the amide group, respectively. 46 Mel-loaded NISVs showed absorption signals at wavelengths of 1651 and 1555 cm −1 , which were slightly shifted from the characteristic signals of Mel. This presumably was due to the interaction between the amide groups of Mel and the NISVs.…”

Analysis of heterogeneity in nonspecific PEGylation reactions of biomolecules

“…Previous reports have shown that it is possible to (at least approximately) determine the composition of PEGylated protein systems using spectrometric techniques such as matrix‐assisted laser desorption ionization time‐of‐flight (MALDI‐TOF) mass spectrometry . In the following, the calculated and experimental distributions of PEG ligands are compared for a number of protein systems with systematically varying N 0 to evaluate the applicability of Eq.…”

Stable Loading and Delivery of Melittin with Lipid-Coated Polymeric Nanoparticles for Effective Tumor Therapy with Negligible Systemic Toxicity

“…Various strategies have been explored to overcome the aforementioned problems, including the alteration of melittin sequence to reduce its hemolysis, PEGylation or glycosylation to increase the stability and prolong the in vivo biological half-life, or the conjugation of targeting molecules, such as the antibody, to increase cancer cell specificity. − However, these methods may decrease the membrane-lytic activity of melittin in cancer cells and have no considerable effect on reducing melittin’s hemolytic activity and tissue toxicity.…”