Analogous copper(I) coordination in metallothionein from yeast and the separate domains of the mammalian protein.
The three-dimensional structures of both vertebrate Cu12-metallothionein (class 1) and yeast Cu8-thionein (class 2) are still unknown. The different copper:protein stoichiometry compared with that of the (ZnCd)7-metallothioneins was expected to alter the metal-thiolate cluster structure considerably. In order to avoid possible domain interactions in the hepatic rat metallothionein, separate chemically synthesized alpha- and beta-domains were used rather than the apoprotein. Apo yeast thionein, and the alpha- and beta-domains of rat liver metallothionein-2 were reconstituted by Cu(I) titration. Reconstitution steps were monitored using spectroscopic methods including luminescence emission and circular dichroism. Upon UV irradiation a linear increase in intensity of the orange-red luminescence was observed near 600 nm up to 6 Cu eq using either compound regardless of the different cysteine sulfer content (yeast thionein 12S, alpha-domain 11S, beta-domain 9S). The characteristic dichroic properties of the yeast copper-protein between 240 and 400 nm were in good agreement with those of the respective class 1 metallothionein domains. All observed Cotton bands were of similar shape and appeared in the same wavelength regions. However, the molar ellipticities were less pronounced in the alpha- and beta-fragments employed. There appears to be a striking similarity between the oligonuclear Cu(I) binding centers in all metallothionein species.
Hartmann
H J
HJ
Anorganische Biochemie, Physiologisch-Chemisches Institut, Universität Tübingen, Germany.
Li
Y J
YJ
Weser
U
U
D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D002942:Circular Dichroism; D003300:Copper / Q000737:chemistry*; D008099:Liver / Q000737:chemistry*; D008668:Metallothionein / Q000737:chemistry*; D051381:Rats; D012441:Saccharomyces cerevisiae / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet
C500769:CUP1-1 protein, S cerevisiae; D002352:Carrier Proteins; C007121:copper thionein; D003300:Copper; D008668:Metallothionein