Evidence for iron channeling in the Fet3p-Ftr1p high-affinity iron uptake complex in the yeast plasma membrane

In high-affinity iron uptake in the yeast Saccharomyces cerevisiae, Fe ^II is oxidized to Fe^III by the multicopper oxidase, Fet3p, and the Fe^III produced is transported into the cell via the iron permease, Ftr1p. These two proteins are likely part of a heterodimeric or higher order complex in the yeast plasma membrane. We provide kinetic evidence that the Fet3p-produced Fe^III is trafficked to Ftr1p for permeation by a classic metabolite channeling mechanism. We examine the ⁵⁹Fe uptake kinetics for a number of complexes containing mutant forms of both Fet3p and Ftr1p and demonstrate that a residue in one protein interacts with one in the other protein along the iron trafficking pathway as would be expected in a channeling process. We show that, as a result of some of these mutations, iron trafficking becomes sensitive to an added Fe^III chelator that inhibits uptake in a strictly competitive manner. This inhibition is not strongly dependent on the chelator strength, however, suggesting that Fe ^III dissociation from the iron uptake complex, if it occurs, is kinetically slow relative to iron permeation. Metabolite channeling is a common feature of multifunctional enzymes. We constructed the analogous ferroxidase, permease chimera and demonstrate that it supports iron uptake with a kinetic pattern consistent with a channeling mechanism. By analogy to the Fe ^III trafficking that leads to the mineralization of the ferritin core, we propose that ferric iron channeling is a conserved feature of iron homeostasis in aerobic organisms.