COVALENT SEQUESTRATION OF THE NITROGEN-MUSTARD MECHLORETHAMINE BY METALLOTHIONEIN

The research reported here demonstrates covalent binding to the metal- binding protein metallothionein (MT) by the therapeutic nitrogen musta rd mechlorethamine. The most surprising aspect of this interaction is the selectivity of the alkylating agent for specific residues of MT. A combination of MS and proteolytic and enzymatic methods was used to d educe specific locations of mechlorethamine alkylation. These experime nts indicated that alkylation occurs predominantly in the carboxyl dom ain of Mi, with one molecule of mechlorethamine covalently cross-linki ng two cysteine residues. Electrospray MS revealed the retention of al l seven metal ions in the cross-linked MT/mechlorethamine adducts, hig hlighting the uniqueness of this protein. Computerized docking experim ents supported the hypothesis that selective binding precedes selectiv e alkylation, and the structure of the drug indicates the minimal stru ctural requirements for this binding. These results support the idea t hat MT overexpressed in tumor cells contributes to the inactivation of anticancer drugs.