Full Length Research Paper

Gangliosides as potential inhibitors of Naja naja venom PLA2 (NV-PLA2) induced human erythrocyte membrane damage

Mukunda Chethankumar and Leela Srinivas*

Adichunchanagiri Biotechnology and Cancer Research Institute, Balagangadharanatha Nagara – 571 448, Karnataka, India.

*Corresponding author. E-mail: leela_srivas@yahoo.co.uk. Tel. +91 – 8234 – 287850/984 Fax: +91 – 8234 – 287984.

Accepted 29 October 2007

   Abstract

We determined the ability of mixed gangliosides (16% GD1b, 19% GT1b, 21% GM1, and 40% GD1a) and individual gangliosides GM1 and GD1b to modulate the NV-PLA₂ induced human erythrocyte ghost membrane damage. CM-Sephadex purification of crude Naja naja venom yielded eight peaks of which peak VII, a major phospholipase A₂ (NV-PLA₂) accounted for 22% of the total protein recovered and 8% of the total PLA₂ activity recovered. The membrane damage induced by NV-PLA₂ was assessed by measuring the decrease in the relative intensity of fluorescence using cis–parinaric acid (PnA) as a monitor molecule. The RBC membranes isolated from healthy human blood showed 72% damage on treatment with NV-PLA₂ (2 mg) when compared to untreated membranes. Mixed gangliosides (18 nM) and GM1 (15 nM) offered 81 and 86% protection respectively, whereas GD1b (20 nM) did not show significant protection. Analysis of membrane bound Na⁺K⁺ and Ca²⁺Mg²⁺ ATPase indicated a 3 fold and 2 folds decrease in their activities on NV-PLA₂ treatment when compared to untreated membranes. Mixed gangliosides restored the Na⁺K⁺ ATPase activity by 78%, whereas GM1 and GD1b offered 74 and 52% restoration respectively. The Ca²⁺Mg²⁺ ATPase activity was restored by 80 and 81% with mixed gangliosides and GM1 respectively. GD1b showed only 50% restoration. Mixed gangliosides and GM1 exhibited a significant dose dependent inhibition of NV-PLA₂ activity when compared to GD1b. The data show that mixed gangliosides and GM1 were effective in modulating NV-PLA₂ induced erythrocyte membrane damage than GD1b.

Key words: Gangliosides, NV-PLA₂, PnA, gangliosides, erythrocyte membrane, Na⁺ K⁺ ATPase, Ca²⁺ Mg²⁺ ATPase.