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. 1991 Jul 15;88(14):6127–6131. doi: 10.1073/pnas.88.14.6127

A copper-thiolate polynuclear cluster in the ACE1 transcription factor.

C T Dameron 1, D R Winge 1, G N George 1, M Sansone 1, S Hu 1, D Hamer 1
PMCID: PMC52035  PMID: 2068093

Abstract

ACE1 is the transcriptional activator of the metallothionein (CUP1 locus) gene in Saccharomyces cerevisiae. Previous data had implicated the N-terminal domain of ACE1 as responsible for the Cu-dependent specific DNA binding. An expression system in Escherichia coli was constructed to enable the isolation of an ACE1 domain containing the DNA and Cu-binding regions. Here we report the purification and characterization of the Cu-ACE1 truncated molecule. Spectroscopic techniques showed that ACE1 contains an unusual type of DNA binding structure that is based on a polynuclear Cu(I)-cysteinyl thiolate cluster. The cluster consists of six or seven Cu(I) ions coordinated to cysteinyl thiolates in a trigonal geometry distorted from planarity. The Cu(I)-cysteine cluster of Cu-ACE1 exhibits structural properties analogous to the Cu(I)-thiolate polynuclear cluster in yeast Cu-metallothionein itself, suggesting an unusual mechanism for the evolution of this regulatory factor. The Cu cluster organizes and stabilizes the conformation of the N-terminal domain of ACE1 for specific DNA binding.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Buchman C., Skroch P., Welch J., Fogel S., Karin M. The CUP2 gene product, regulator of yeast metallothionein expression, is a copper-activated DNA-binding protein. Mol Cell Biol. 1989 Sep;9(9):4091–4095. doi: 10.1128/mcb.9.9.4091. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Byrd J., Berger R. M., McMillin D. R., Wright C. F., Hamer D., Winge D. R. Characterization of the copper-thiolate cluster in yeast metallothionein and two truncated mutants. J Biol Chem. 1988 May 15;263(14):6688–6694. [PubMed] [Google Scholar]
  3. Fürst P., Hamer D. Cooperative activation of a eukaryotic transcription factor: interaction between Cu(I) and yeast ACE1 protein. Proc Natl Acad Sci U S A. 1989 Jul;86(14):5267–5271. doi: 10.1073/pnas.86.14.5267. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Fürst P., Hu S., Hackett R., Hamer D. Copper activates metallothionein gene transcription by altering the conformation of a specific DNA binding protein. Cell. 1988 Nov 18;55(4):705–717. doi: 10.1016/0092-8674(88)90229-2. [DOI] [PubMed] [Google Scholar]
  5. George G. N., Byrd J., Winge D. R. X-ray absorption studies of yeast copper metallothionein. J Biol Chem. 1988 Jun 15;263(17):8199–8203. [PubMed] [Google Scholar]
  6. George G. N., Winge D., Stout C. D., Cramer S. P. X-ray absorption studies of the copper-beta domain of rat liver metallothionein. J Inorg Biochem. 1986 Jul;27(3):213–220. doi: 10.1016/0162-0134(86)80062-9. [DOI] [PubMed] [Google Scholar]
  7. Good M., Hollenstein R., Sadler P. J., Vasák M. 113Cd NMR studies on metal-thiolate cluster formation in rabbit Cd(II)-metallothionein: evidence for a pH dependence. Biochemistry. 1988 Sep 6;27(18):7163–7166. doi: 10.1021/bi00418a074. [DOI] [PubMed] [Google Scholar]
  8. Grassetti D. R., Murray J. F., Jr Determination of sulfhydryl groups with 2,2'- or 4,4'-dithiodipyridine. Arch Biochem Biophys. 1967 Mar;119(1):41–49. doi: 10.1016/0003-9861(67)90426-2. [DOI] [PubMed] [Google Scholar]
  9. Hu S., Fürst P., Hamer D. The DNA and Cu binding functions of ACE1 are interdigitated within a single domain. New Biol. 1990 Jun;2(6):544–555. [PubMed] [Google Scholar]
  10. Struhl K. Helix-turn-helix, zinc-finger, and leucine-zipper motifs for eukaryotic transcriptional regulatory proteins. Trends Biochem Sci. 1989 Apr;14(4):137–140. doi: 10.1016/0968-0004(89)90145-X. [DOI] [PubMed] [Google Scholar]
  11. Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
  12. Szczypka M. S., Thiele D. J. A cysteine-rich nuclear protein activates yeast metallothionein gene transcription. Mol Cell Biol. 1989 Feb;9(2):421–429. doi: 10.1128/mcb.9.2.421. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Welch J., Fogel S., Buchman C., Karin M. The CUP2 gene product regulates the expression of the CUP1 gene, coding for yeast metallothionein. EMBO J. 1989 Jan;8(1):255–260. doi: 10.1002/j.1460-2075.1989.tb03371.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Winge D. R., Nielson K. B., Gray W. R., Hamer D. H. Yeast metallothionein. Sequence and metal-binding properties. J Biol Chem. 1985 Nov 25;260(27):14464–14470. [PubMed] [Google Scholar]

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