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Evidence for pH Dependent Zn2+influx in K562 Erythroleukemia Cells: Studies Using ZnAF-2F Fluorescence and 65Zn2+ Uptake

Evidence for pH Dependent Zn2+influx in K562 Erythroleukemia Cells: Studies Using ZnAF-2F Fluorescence and 65Zn2+ Uptake

Robert A Colvin, Charles P Fontaine, Dustin Thomas, Tomoya Hirano, Tetsuo Nagano, Kazuya Kikuchi

Arch Biochem Biophys. 2005 Oct 15;442(2):222-8.

PMID: 16183033

Abstract:

Using both ZnAF-2F (a Zn2+ specific fluorophore) and 65Zn2+, we determined the rate of transporter mediated Zn2+ influx (presumably mediated by the SLC39A1 gene product, protein name hZIP1) under steady state conditions and studied the effects of extracellular acidification. When K562 erythroleukemia cells were placed in Zn2+ containing buffers (1-60 microM), the initial rate of 65Zn2+ accumulation mirrored the apparent rise in free intracellular Zn2+ concentrations sensed by ZnAF-2F. Therefore, newly transported Zn2+ equilibrated with the free intracellular Zn2+ pool sensed by ZnAF-2F. A new steady state with elevated free intracellular Zn2+ was established after about 30 min. An estimate of 11 microM for the Km and 0.203 nmol/mg/s for the Vmax were obtained for Zn2+ influx. 65Zn2+ uptake and ZnAF-2F fluorescent changes were inhibited by extracellular acidification (range tested: pH 8-6, IC50 = pH 6.34). The IC50 for proton effects was close to the pKa for histidine, suggesting conserved histidine residues present in SLC39A1 play a critical role in Zn2+ influx and are involved in the pH effect.

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
AP443302098	ZnAF-2F		443302-09-8	Price

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