Proteomic and glycomic analyses of venom geographic variations of Wagler's pit vipers and king cobra
Date Issued
2012
Date
2012
Author(s)
Chang, Hui-Ching
Abstract
Herein, we have investigated the geographic variations of Tropidolaemus wagleri from two regions and those of O. hannah from four regions by venom protein purifications, identification and biofuntional characterizations using the proteomic and glycomic approaches.
Part I. Phospholipases A2 (PLA2s) were purified and characterized from Tropidolaemus wagleri venoms collected from Sulawesi and Sumatra. An active PLA2 and an inactive Lys49-like PLA2 could be isolated from each venom sample. Mass analyses and N-terminal sequences revealed that the PLA2 variants from the two venom samples were different although their neurotoxic waglerins were identical. Thus, the samples were probably derived from two species (T. subannulatus and T. wagleri), consistent with the recent taxonomic study of this genus. Since the Glu6 residue was conserved in both the PLA2s, the PLA2s from Sulawesi and Sumatra were designated as Tsu-E6 and Twa-E6, respectively. Interestingly, both Tsu-E6 and Twa-E6 appeared to be glycosylated at Asn14. Hydrolysis by PNGase F reduced their apparent masses from 16 to 14 kDa. The released glycans
were further analyzed by MALDI-TOF and shown to be complex type oligosaccharides without sialylation. Tsu-E6 inhibited ADP-induced aggregation of mouse and human platelets, and Twa-E6 inhibited the ADP- or collagen-induced aggregation of human platelets, but stimulated the aggregation of mouse platelets. However, enzymatic removal of glycans from both the PLA2s did not significantly alter their effects on lipid hydrolysis and platelet aggregation. The thermostability
of Twa-E6 was found to be as good as those of other homologous PLA2s. The presence of these oligosaccharides on snake venom PLA2s therefore warrants further analyses.
Part II. We profiled and analyzed four geographic venom samples of O. hannah which were obtained from Indonesia, Malaysia, Guangxi, and Hai-Nan. The enzyme activities, toxicities on mice of the four venoms were investigated. The PLA2s and 3FTxs were purified. There are great geographic variations in the enzyme content, protein composition and toxic properties between the Southeast Asian and Chinese king cobra venoms. The OH-APLA2 is the major PLA2 conserved in the venoms from China, and the venom from Indonesia and Malaysia express a PLA2 similar to OH-APLA2 II as judged by PMF analysis. Chinese king cobra venoms appear to have lower
metalloprotease or hemorrahagin activities but higher prothrombinase activities. Even though the oh-55 3FTx is conserved in king cobra venoms, it is not the most abundant in all the kign cobra venom. The composition of the other 3FTx variants is highly variable. Finally, the venom from
Indonesia is especially performing high alkaline phosphomonoesterase activity, which may present synergistic reaction in the toxic property of venom. This will need a further investigation to resovle.
Subjects
geographic variation
venom phospholipases A2
glycan structure
platelet aggregation inhibitor
Tropidolaemus wagleri
Ophiophagus hannah
three-finger toxin
Type
thesis
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