Sequential 1H-NMR assignments of mouse
[Cd7]-metallothionein-1 (MT1) have
been carried out by standard homonuclear NMR methods and
the use of an accordion–heteronuclear multiple quantum
correlation (HMQC) experiment for establishing the metal,
113Cd2+, to cysteine connectivities.
The three-dimensional structure was then calculated using
the distance constraints from two-dimensional nuclear Overhauser
effect (NOE) spectroscopy spectra and the Cys–Cd
connectivities as input for a distance geometry-dynamical
simulated annealing protocol in X-PLOR 3.851. Similar to
the mammalian MT2 isoforms, the homologous primary structure
of MT1 suggested two separate domains, each containing
one metal cluster. Because there were no interdomain constraints,
the structure calculation for the N-terminal β- and
the C-terminal α-domain were carried out separately.
The structures are based on 409 NMR constraints, consisting
of 381 NOEs and 28 cysteine–metal connectivities.
The only elements of regular secondary structure found
were two short stretches of 310 helices along
with some half-turns in the α-domain. Structural comparison
with rat liver MT2 showed high similarity, with the β-domain
structure in mouse MT1 showing evidence of increased flexibility
compared to the same domain in MT2. The latter was reflected
by the presence of fewer interresidue NOEs, no slowly exchanging
backbone amide protons, and enhanced cadmium–cadmium
exchange rates found in the β-domain of MT1.