HgEDTA complex inhibits GTP interactions with the E-site of brain beta-tubulin.

We have found that EDTA and EGTA complexes of Hg2+, which conventional wisdom has assumed are biologically inert, are potentially injurious to the neuronal cytoskeleton. Tubulin, a major protein component of the neuronal cytoskeleton, is the target of multiple toxicants, including many heavy metal ions. Among the mercurials, inorganic mercuric ion (Hg2+) is one of the most potent inhibitors of microtubule polymerization both in vivo and in vitro. In contrast to other heavy metals, the capacity of Hg2+ to inhibit microtubule polymerization or disrupt formed microtubules cannot be prevented by the addition of EDTA and EGTA, both of which bind Hg2+ with very high affinity. To the contrary, the addition of these two chelating agents potentiates Hg2+ inhibition of tubulin polymerization. Results herein show that HgEDTA and HgEGTA inhibit tubulin polymerization by disrupting the interaction of GTP with the E-site of brain beta-tubulin, an obligatory step in the polymerization of tubulin. Both HgEDTA and HgEGTA, but not free Hg2+, prevented binding of [32P]8N3GTP, a photoaffinity nucleotide analog of GTP, to the E-site and displaced bound [32P]8N3GTP at low micromolar concentrations. This complete inhibition of photoinsertion into the E-site occurred in a concentration- and time-dependent fashion and was specific for Hg2+ complexes of EDTA and EGTA, among the chelating agents tested. Given the ubiquity of Hg2+ in the environment and the widespread use of EDTA in foodstuffs and medicine, these mercury complexes may pose a potentially serious threat to human health and play a role in diseases of the neuronal cytoskeleton.