MERCURY BINDING TO METALLOTHIONEINS - FORMATION OF THE HG18-MT SPECIES

We report the formation and properties of a novel mercury-protein complex formed from rabbit liver metallothionein, namely Hg18-MT. Complex formation is dependent on the presence of the Cl- ion and a pH below 6. Rabbit liver Hg18-MT exhibits a strongly dichroic CD signal in the 240-360-nm region, and a unique X-ray absorption, near-edge spectrum (Lu, W.; Kasrai, M.; Bancroft, G. M.; Stillman, M. J.; Tan, K. H. Inorg. Chem. 1990, 29, 2561-2563). Hg18-MT elutes with an apparent molar mass of 5500 Da from Sephadex G-50 columns. There is no indication of dimer formation. Formation of the 3-dimensional structure associated with Hg18-MT is unique to rabbit liver isoform 2; unless isoform 1 is initially lyophilized and redissolved in acidic solution, the CD spectrum characteristic of Hg18-MT is not observed. The CD spectrum of Hg18-MT can be used to discriminate between isoforms 1 and 2 following chromatographic separation on Sephadex-DEAE columns. Isoforms of rat metallothionein do not adopt the same structure as isoforms of rabbit metallothionein. Neither isoform of metallothionein isolated from rat liver binds 18 Hg2+ to form a complex characterized by a similar CD signal. Each of the four MT isoforms studied binds 7 mol of Hg2+ to form Hg7-MT species that exhibit similar CD spectra at pH 7. The CD bands observed for the Hg-containing metallothioneins lie under bands assigned generally as sulfur-to-mercury charge transfer, although the strongly dichroic band of Hg18-MT 2 lies to the red of bands previously reported for other Hg-MT complexes. We suggest that the CD spectral intensity observed for rabbit liver Hg18-MT arises from the enhanced chirality of stacked Hg-S bonds as the peptide chain self-associates into a unique 3-dimensional structure based on a single domain. This structure represents a third structural motif for metallothioneins. The essential requirement for Cl- suggests that the coordination geometry of the Hg2+ may be pseudotetrahedral with bridging thiolates and outlying chlorides. This would result in a kink in the structure which could be the origin of the enhanced CD intensity. We propose that the significant differences in complexation properties observed for the different isoforms result from differences in the amino acid sequences between each of the MT isoforms in the region of a pleat required by the Hg18-MT structure.