Binding of excess cadmium(II) to Cd<inf>7</inf>-metallothionein from recombinant mouse Zn<inf>7</inf>-metallothionein 1. UV-VIS absorption and circular dichroism studies and theoretical location approach by surface accessibility analysis

Neus Cols, Núria Romero-Isart, Mercè Capdevila, Baldomero Oliva, Pilar Gonzàlez-Duarte, Roser Gonzàlez-Duarte, Sìlvia Atrian

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Abstract

A mouse metallothionein (MT) 1 expression system has been constructed that renders recombinant MT as a high purity Zn-coordinated protein. Spectral changes in absorption and circular dichroism following the addition of up to 7 tool equivalents of Cd2+ to recombinant Zn7-MT showed that it behaves like the native protein. Exposure of Cd7-MT to Cd2+ resulted in further binding of these ions to the protein, although saturation was not achieved on the addition of up to 22 tool equivalents of Cd2+ to Zn7-MT. Spectral data are compatible with a model in which the first four additional Cd2+ ions are bound to Cd7-MT via sulfur atoms, and indicate that no further thiol groups are involved in the binding of the excess Cd(II) over 11. Cd2+ ions bound in excess to Cd7-MT appear to have lower binding constants as exposure of Cd(n)-MT (n > 7) species to Chelex-100 retrieved Cd7-MT. Based on the X- ray data, the accessible surface areas of sulfur atoms in Cd5,Zn2-MT 2 were calculated. This led us to propose that the coordination of the first three additional Cd(II) ions to Cd7-MT proceeds by means of S-Met1-O-Met1, S- Cys7-S-Cys13 and S-Cys5-SCys26 pairs. Finally, comparison of the behavior of the entire MT with that of the recombinant α MT and β MT subunits indicates that mutual influences may not be negligible.
Original languageEnglish
Pages (from-to)157-166
JournalJournal of Inorganic Biochemistry
Volume68
DOIs
Publication statusPublished - 15 Nov 1997

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