Abstract
BackgroundSnake venoms are a complex mixture of active principles mainly peptides and proteins also including amino acids, nucleotides, free lipids, carbohydrates and metallic elements bound to proteins that interfere in several biological systems. In this study, we aimed to understand the mode of action of the apoptosis inducing ability of Naja naja venom phospholipase A2 (NV-PLA2) using isolated human peripheral lymphocytes.ResultsHuman peripheral lymphocytes when incubated with Naja naja venom phospholipase A2 (NV-PLA2) induced up to 68% DNA fragmentation. The dialysed conditioned media obtained by incubating lymphocytes with NV-PLA2 at 15th min induced 44% DNA fragmentation, referred to as cmlp-active. Cmlp-active showed 20.5% increased protein concentration than the corresponding control condition media cmlp-c-15. Test for creatine kinase activity in cmlp-active proved negative and negligible amount of lactate dehydrogenase did not show significant DNA fragmentation. Fractionation of cmlp-active on Sephadex G-25 showed two peaks, major peak induced 38% DNA fragmentation, which was further rechromatographed on Sephadex G-25. The single peak obtained was named PID15 (Phospholipase A 2 Induced DNA fragmentation factor secreted at 15 th min). Q-Tof MS/MS analysis of PID-15 showed it is a 6 kDa peptide. PID15 sequence analysis gave 40 amino acids in the following order, msilpcknvs iwvikdtaas dkevvlgsdr aikflylatg. The homology search for the sequence revealed it to be an Apoptosis Inducing Factor (AIF).ConclusionResults indicate that the secretion of PID15 is dependent on concentration of NV-PLA2 treatment, incubation time and also on temperature and the probable membrane origin of PID15 and not of cytosolic origin with apoptosis inducing ability.
Highlights
Snake venoms are a complex mixture of active principles mainly peptides and proteins including amino acids, nucleotides, free lipids, carbohydrates and metallic elements bound to proteins that interfere in several biological systems [6,7]
Venom phospholipase A2 (PLA2) are able to induce apoptosis through calpain mediated activation of caspases and other pro apoptotic proteins, and by formation of permeability transition pore in the membrane as a result of increased Ca2+ in the mitochondrial matrix [18]
Earlier studies have shown that a cytotoxic substance from Korean snake venom which is responsible for the apoptosis is L-amino acid oxidase, a glycoprotein of 110 kDa
Summary
Snake venoms are a complex mixture of active principles mainly peptides and proteins including amino acids, nucleotides, free lipids, carbohydrates and metallic elements bound to proteins that interfere in several biological systems. 450 of the 3000 species of snakes found worldwide are considered to be dangerous to humans. Of these about 52 are encountered in the geographical area, which extends from Pakistan and the rest of the Indian subcontinent through to the Philippines and Indonesia. Symptoms of snake envenomation are a combination of the complex protein and non protein components present in the venom. These effects at later stages result in tissue damage, which reaches many organs, such as the brain, lung, kidney, heart, and liver [11,12,13]
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