Metal-response element-binding transcription factor-1 complex with MRE DNA

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NMR assignment of the six zinc fingers of MTF-1 in the free and DNA-bound states.

Metallothioneins (MTs) are small cysteine-rich proteins whose structure is
conserved from fungi to man. MTs strongly bind heavy metals, notably zinc,
copper and cadmium. Upon exposure of cells to heavy metal and other adverse
treatments, MT gene transcription is strongly enhanced. Metal induction is
mediated by several copies of a 15 bp consensus sequence (metal-responsive
element, MRE) present in the promoter region of MT genes. We and others have
demonstrated the presence of an MRE-binding factor in HeLa cell nuclear
extracts. We found that this factor, termed MTF-1 (MRE-binding transcription
factor) is inactivated/reactivated in vitro by zinc withdrawal/addition. Here
we report that the amounts of MTF-1-DNA complexes are elevated several-fold in
zinc-treated cells, as measured by bandshift assay. We have also cloned the
cDNA of mouse MTF-1, a 72.5 kDa protein. MTF-1 contains six zinc fingers and
separate transcriptional activation domains with high contents of acidic and
proline residues. Ectopic expression of MTF-1 in primate or rodent cells
strongly enhances transcription of a reporter gene that is driven by four
consensus MREd sites, or by the complete mouse MT-I promoter, even at normal
zinc levels.

Radtke, F., Heuchel, R., Georgiev, O., Hergersberg, M., Gariglio, M., Dembic,
Z., and Schaffner, W. (1993). Cloned transcription factor MTF-1 activates the
mouse metallothionein I promoter. EMBO J., 12, 1355-1362.

Cloned transcription factor MTF-1 activates the mouse metallothionein I promoter.

metal-response element-binding transcription factor-1

1H, 13C and 15N resonance assignments for the six-zinc finger domain of the
MTF-1 metalloregulatory protein in the DNA bound state

A cryogenically cooled HCN Triax probe (cryoprobe) was used for all experiments

update DNA residue label to two-letter code

All spectra were recorded with 95% H2O/5% D2O

Protein was produced in a reduced lyopholized form (free thiols). Protein was subsequently resuspended in degassed buffer under anaerobic conditions. Protein was titrated into DNA solution in small aliquots to make complex.

Delaglio, F., Grzesiek, S., Vuister, G.W., Zhu, G., Pfeifer, J., & Bax, A.
(1995). NMRPipe: a multidimensional spectral processing system based on UNIX
pipes. J. Biomol. NMR 6, 277-293.

Johnson, B.A. and Blevins, R.A.(1994) NMRView: A computer program for the
visualization and analysis of NMR Data. J. Biomol. NMR 4, 603-614.