Lipolytic activity of ricin from Ricinus sanguineus and Ricinus communis on neutral lipids Available to Purchase

The present study was carried out with a view of determining ricin lipolytic activity on neutral lipids in emulsion and in a membrane-like model. Using 2,3-dimercapto-1-propanol tributyrate (BAL-TC₄) as substrate, the lipolytic activity of ricin was found to be proportional to ricin and substrate concentrations, with an apparent K_m (K_m,app) of 2.4mM, a k_cat of 200min⁻¹ and a specific activity of 1.0unit/mg of protein. This work was extended to p-nitrophenyl (pNP) fatty acid esters containing two to twelve carbon atoms. Maximum lipolytic activity was registered on pNP decanoate (pNPC₁₀), with a K_m,app of 3.5mM, a k_cat of 173min⁻¹ and a specific activity of 3.5units/mg of protein. Ricin lipolytic activity is pH and galactose dependent, with a maximum at pH7.0 in the presence of 0.2M galactose. Using the monolayer technique with dicaprin as substrate, ricin showed a lipolytic activity proportional to the ricin concentration at 20mN/m, which is dependent on the surface pressure of the lipid monolayer and is detectable up to 30mN/m, a surface pressure that is of the same order of magnitude as that of natural cell membranes. The methods based on pNPC₁₀ and BAL-TC₄ hydrolysis are simple and reproducible; thus they can be used for routine studies of ricin lipolytic activity. Ricin from Ricinus communis and R. sanguineus were treated with diethyl p-nitrophenylphosphate, an irreversible serine esterase inhibitor, and their lipolytic activities on BAL-TC₄ and pNPC₁₀, and cytotoxic activity, were concurrently recorded. A reduction in lipolytic activity was accompanied by a decrease in cytotoxicity on Caco2 cells. These data support the idea that the lipolytic activity associated with ricin is relevant to a lipase whose activity is pH and galactose dependent, sensitive to diethyl p-nitrophenylphosphate, and that a lipolytic step may be involved in the process of cell poisoning by ricin. Both colorimetric tests used in this study are sensitive enough to be helpful in the detection of possible lipolytic activities associated with other cytotoxins or lectins.