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PEPTIDE BINDING AS A MODE OF ACTION FOR THE CARCINOGENICITY AND TOXICITY OF ARSENIC
Citation:
KITCHIN, K. T. AND K. WALLACE. PEPTIDE BINDING AS A MODE OF ACTION FOR THE CARCINOGENICITY AND TOXICITY OF ARSENIC. Presented at 1st Georgian Bay International Conference on Bioinorganic Chemistry, Parry Sound, ON, CANADA, May 22 - 25, 2007.
Description:
Arsenic exposure leads to tumors in human skin, lung, urinary bladder, kidney and liver. Three likely initial stages of arsenical-macromolecular interaction are (1) binding of trivalent arsenicals to the sulfhydryl groups of peptides and proteins, (2) arsenical-induced generation of free radicals and oxidative stress and (3) altered DNA methylation status. The binding of trivalent arsenicals to the sulfllydryl groups of peptides and proteins can be an important component of several of the proposed modes of action for arsenic - chromosomal abnormalities, altered DNA repair, DNA methylation changes and enhanced cell proliferation dependent pathways (both stimulatory and necrosis induced). Recent binding studies have shown that arsenite binds to the four cysteine containing zinc finger region of the human estrogen receptor-alpha [sequence of RYCAVCNDYASGYHYGVWSCEGCKA] with a Kd of 2.2 µMolar. If the final three cysteines are deleted from the sequence, the Kd increases to 190 µMolar. Binding studies were performed with peptides containing two cysteines separated by 0 to 17 intervening amino acids. Increasing the number of intervening amino acids from 0 to 14 made little difference in the observed Kd values for arsenite (2.7 to 20.1 µMolar). With 19 intervening amino acids, the Kd value increased to 123. µMolar. These binding data suggest that the studied shorter peptides are flexible enough to form dithiol binding sites for arsenite. Arsenite forms complexes of vastly different stabilities with peptides (either inter- or intra- molecularly) when it binds in a mono-, bi-, or tri- dentate fashion. These arsenite-peptide complexes have half lives of about < 1 second, 1 minute and 2 hours, respectively. A mathematical model of arsenite binding simultaneously to cellular monothiol sites and dithiol sites has been created. This binding model predicts that in the range of 1 nanoMolar to 1 milliMolar total arsenite concentration: (a) 99% of the arsenite is bound to sulfhydryl groups and (b) 90 to 98% of the total arsenite binding is to monothiol sites. At 10 µMolar total arsenite concentration, the model predicts that only 0.075% of the monothiol binding sites are occupied by arsenite and that 4.3% of the dithiol sites are occupied by arsenite. Thus, low Kd dithiol or trithiol biological binding sites are excellent candidates for causing the adverse biological effects of arsenic exposure, particularly if the mode of action is by altered protein function (e.g. incorrect DNA repair) rather than by enzyme inhibition. Future studies with 14-C labeled forms of monomethylarsonous acid and dimethylarsinous acid will provide new binding data that compliments existing data with inorganic radioactive 73-As arsenite and lead to a comprehensive model of how important trivalent arsenicals interact with cysteine containing peptides and proteins in biological systems.