Structural analysis of phospholipase A2 by near-IR Fourier transform Raman spectroscopy

Venom toxins were isolated from Formosan cobra (Naja naja atra) by cation-exchange chromatography. Most toxin components could be obtained in relatively pure forms by single-step ion-exchange chromatography whereas an extra step of gel permeation was needed for the separation of phospholipase A2 (PLA2) from the major neurotoxic component, i.e. cobrotoxin. The newer near-IR FT-Raman analytical method has been applied to the characterization of PLA2 in their lyophilized forms. Structural analysis of PLA2 and correlation of Raman spectroscopic data with amino acid compositions were made. The results indicate that phospholipase A2 showed the Raman peak at 1659 cm-1 which is characteristic of the alpha-helical structure in this enzyme. It is also found that the relative Raman signal intensities of Tyr, Phe, Trp and Met residues in purified toxins correlate very well with the structural data obtained from amino acid analysis. The application of near-IR FT-Raman techniques in the detection of the microenvironments of the aromatic amino acids such as Tyr and Trp in the native toxins may prove useful in the investigation of the functional properties of various venom toxins.