Inhibition of tubulin guanosine-5′-triphosphatase by lipid peroxides: Protective effects of vitamin A derivatives

Lipid peroxidation of cellular proteins has been postulated to be involved in cellular aging. Given the importance of the cytoskeleton in cellular function, it is a prime candidate as a potential target for the deleterious effects of lipid peroxides. In this study, the effects of lipid peroxides on microtubule assembly have been studied in an in vitro assay system, as have the protective effects of the vitamin A group (β-carotene, retinal, and retinol). The assay was based on tubulin guanosine-5′-triphosphatase (GTPase) activity, which is associated with all steps of microtubule assembly. Soybean lecithin was utilized as the starting point to generate lipid peroxides. Its selection was based on the high proportion of phospholipids in the cellular membrane. Lipid peroxides were generated by photooxidation of lecithin, dissolved in methanol, in the presence of 0.004% methylene blue at 4°C for 8 h. Lipid peroxides (1.0 mg/mL) inhibited tubulin GTPase by 49%, relative to the control. Vitamin A derivatives (retinol, retinal, and β-carotene) all had the ability to protect against the inhibitory effects of lipid peroxides, presumably owing to their antioxidant activities. This protective effect was more pronounced when utilizing a 30-min, as opposed to a 15-min, reaction time. This suggests a relatively slow rate of reaction between the peroxide and vitamin A group. These studies present a mechanism for the ability of vitamin A to inhibit aging of the cell.