Kinetic identification of a mitochondrial zinc uptake transport process in prostate cells

Abstract

Prostate cells accumulate high cellular and mitochondrial concentrations of zinc, generally 3-10-fold higher than other mammalian cells. However, the mechanism of mitochondrial import and accumulation of zinc from cytosolic sources of zinc has not been established for these cells or for any mammalian cells. Since the cytosolic concentration of free Zn(2+) ions is negligible (estimates vary from 10(-9) to 10(-15) M), we postulated that loosely bound zinc-ligand complexes (Zn-Ligands) serve as zinc donor sources for mitochondrial import. Zinc chelated with citrate (Zn-Cit) is a major form of zinc in prostate and represents an important potential cytosolic source of transportable zinc into mitochondria. The mitochondrial uptake transport of zinc was studied with isolated mitochondrial preparations obtained from rat ventral prostate. The uptake rates of zinc from Zn-Ligands (citrate, aspartate, histidine, cysteine) and from ZnCl(2) (free Zn(2+)) were essentially the same. No zinc uptake occurred from either Zn-EDTA, or Zn-EGTA. Zinc uptake exhibited Michaelis-Menten kinetics and characteristics of a functional energy-independent facilitative transporter associated with the mitochondrial inner membrane. The uptake and accumulation of zinc from various Zn-Ligand preparations with logK(f) (formation constant) values less than 11 was the same as for ZnCl(2;) and was dependent upon the total zinc concentration independent of the free Zn(2+) ion concentration. Zn-Ligands with logK(f) values greater than 11 were not zinc donors. Therefore the putative zinc transporter exhibits an effective logK(f) of approximately 11 and involves a direct exchange of zinc from Zn-Ligand to transporter. The uptake of zinc by liver mitochondria exhibited transport kinetics similar to prostate mitochondria. The results demonstrate the existence of a mitochondrial zinc uptake transporter that exists for the import of zinc from cytosolic Zn-Ligands. This provides the mechanism for mitochondrial zinc accumulation from the cytosol which contains a negligible concentration of free Zn(2+). The uniquely high accumulation of mitochondrial zinc in prostate cells appears to be due to their high cytosolic level of zinc-transportable ligands, particularly Zn-Cit.

Terminal oxidation and the effects of zinc in prostate versus liver mitochondria

“…This reveals that the inhibitory effect involves a direct intermolecular transfer of zinc from the donor ZnLigand to the recipient electron transport component. A similar mechanism is involved in the import of zinc into mitochondria by a putative inner membrane zinc uptake transporter that has an apparent log K f ~ 11 (Guan et al, 2003). Consistent with this mechanism, Halthout et al (2001) reported that the direct intermolecular transfer of zinc from donor ligand to recipient peptide does not involve an intermediary free Zn + + ion step, and is dependent upon the relative zincbinding affinities of the interacting components.…”

“…The ZnLigand solutions contained 20 μM ZnCl 2 and 60 μM ligand. The basis for these concentrations and characterizations of zinc ligand forms and their reactivity is presented in Guan et al (2003). The log K f values (formation constants) are ZnCitrate ~ 5, ZnCysteine ~ 10, ZnEDTA ~ 16.…”

Zinc as an anti-tumor agent in prostate cancer and in other cancers

“…Therefore the cytosolic source of mitochondrial zinc is derived from mobile reactive ZnLigands that are 10 kDa or less and are capable of permeating the outer mitochondrial membrane pores. These low molecular weight ZnLigands (such as ZnMetallothionein, ZnCitrate, ZnAspartate, ZnHistidine) constitute the source of "mobile reactive zinc" for transport into the mitochondria [14][15][16][17][18]. This pool of zinc is probably in the low uM range.…”

“…This pool of zinc is probably in the low uM range. A mitochondrial zinc uptake transporter activity that is capable of importing zinc from these cytosolic ZnLigands has been kinetically identified in prostate and liver mitochondria [17,18]. The putative transporter effectively transports zinc from ZnLigands that have a binding affinity lower than about log Kf=12.…”

Mitochondrial function, zinc, and intermediary metabolism relationships in normal prostate and prostate cancer

“…In addition, ZnMT (logKf~10) is an effective donor for zinc uptake by mitochondria, and this might be an important zinc chaperone in cells that have high expression of MT and low levels of other mobile reactive ZnLigands such as liver cells (Costello et al, 2004b). This zinc uptake process involves a kinetically-identified zinc uptake transporter that is associated with the inner mitochondrial membrane (Guan et al, 2003). The mechanism involves the direct intermolecular exchange (as described above) of zinc from the donor ZnLigand to the recipient putative uptake transporter protein which has an apparent logKf~10-11.…”