Structure-function analyses of the ATX1 metallochaperone

Saccharomyces cerevisiae Atx1p represents a member of the family of metallo chaperone molecules that escort copper to distinct intracellular targets, A tx1p specifically delivers copper to the Ccc2p copper transporter in the Go lgi. Additionally, when overproduced, Atx1p substitutes for superoxide dism utase 1 in preventing oxidative damage; however the mechanistic overlap bet ween these functions is unresolved. The crystal structure of Atx1p has been solved recently, By examining a surface electrostatic potential distributi on, multiple conserved lysines are revealed on one face of Atx1p. An additi onal conserved lysine (Lys(65)) lies in close proximity to the metal bindin g site. Through site-directed mutagenesis, residues in the metal binding re gion including Lys65 were found to be necessary for both copper delivery to Ccc2p and for Atx1p antioxidant activity. Copper trafficking to Ccc2p also relied on the lysine rich face of Atx1p. Surprisingly however, elimination of these lysines did not inhibit the antioxidant activity of Atx1p, We pro vide evidence that Atx1p does not suppress oxidative damage by a metallocha perone mechanism but may directly consume superoxide. Purified Cu-Atx1p rea cts noncatalytically with superoxide anion in vitro. We conclude that the c opper-trafficking and antioxidant functions of Atx1p arise from chemically and structurally distinct attributes of this metallochaperone.