A copper-thiolate polynuclear cluster in the ACE1 transcription factor.
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.
Dameron
C T
CT
University of Utah Medical Center, Salt Lake City 84132.
Winge
D R
DR
George
G N
GN
Sansone
M
M
Hu
S
S
Hamer
D
D
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D003300:Copper / Q000032:analysis*; D003545:Cysteine / Q000032:analysis*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D004577:Electron Probe Microanalysis; D004926:Escherichia coli / Q000235:genetics; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D013056:Spectrophotometry, Ultraviolet; D014157:Transcription Factors / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism
C060256:CUP2 protein, S cerevisiae; D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D003300:Copper; D003545:Cysteine