Summary
Zn2+ has been allowed to equilibrate across the red cell membrane using two agents that increase membrane permeability to this ion: the ionophore A23187 and the specific carrier ethylmaltol. Extracellular free Zn2+ was controlled with EGTA (1,2-di(2-aminoethoxy)ethane-NNN′N′tetra-acetic acid)) buffers, except in the case of ethylmaltol, which itself acts as a buffer. Measurement of cellular zinc content at different levels of free Zn2+ facilitated the study of intracellular Zn2+ binding. It was also possible to estimate intracellular free Zn2+ concentration in untreated cells using a “null-point” technique. Intracellular zinc was found to consist of an inexchangeable component of about 129 μmol/1013 cells and an exchangeable component of 6.7±1.5 μmol/1013 cells, with a free concentration of about 2.4×10−11 m. The main component of Zn2+ buffering is hemoglobin, with a dissociation constant of about 2×10−8 m.
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References
Bettger, W.J., Taylor, C.G. 1986. Effects of copper and zinc status of rats on the concentration of copper and zinc in the erythrocyte membrane.Nutr. Res. 6:451–457
Ferreira, H.G., Lew, V.L. 1976. Use of ionophore A23187 to measure cytoplasmic Ca buffering and activation of the Ca pump by internal Ca.Nature 259:47–49
Flatman, P.W., Lew, V.L. 1980. Magnesium buffering in intact human red blood cells measured using the ionophore A23187.J. Physiol. 305:13–30
Foote, J.W., Delves, H.T. 1984. Albumin bound and α2-macroglubolin bound zinc concentrations in the sera of healthy adults.J. Clin. Pathol. 37:1050–1054
Galdes, A., Vallee, B.L. 1983. Categories of zinc metalloenzymes.In: Metal Ions in Biological Systems. H. Sigel, editor.15:1–54, Marcel Dekker, New York
Gilman, J.G., Brewer, G.J. 1978. The oxygen-linked zinc-binding site of human haemoglobin.Biochem. J. 169:625–632
Giroux, E.L., Henkin, R.I. 1972. Competition for zinc among serum albumin and amino acids.Biochim. Biophys. Acta 273:64–72
Grider, A., Bailey, L.B., Cousins, R.J. 1990. Erythrocyte metallothionein as an index of zinc status in humans.Proc. Natl. Acad. Sci. USA 87:1259–1262
Harris, W.R., Keen, C. 1989. Calculations of the distribution of zinc in a computer model of human serum.J. Nutr. 119:1677–1682
Hider, R.C., Ejim, L., Taylor, P.D., Gale, R., Huehms, E., Porter, J. 1990. Facilitated uptake of zinc into human erythrocytes. Relevance to the treatment of sickle-cell anaemia.Biochem. Pharmacol. 39:1005–1012
Kalfakakou, V., Simons, T.J.B. 1990. Anionic mechanisms of zinc uptake across the human red cell membrane.J. Physiol. 421:485–497
Magneson, G.R., Puvathingal, J.M., Ray, W.J. 1987. The concentrations of free Mg2+ and free Zn2+ in equine blood plasma.J. Biol. Chem. 262:11140–11148
Martell, A.E., Smith, R.M. 1974. Critical Stability Constants: Amino Acids. Vol. 1. Plenum, New York
Martell, A.E., Smith, R.M. 1982. Critical Stability Constants: First Supplement. Vol. 5. Plenum, New York
Ohno, H., Doi, R., Yamamura, K., Yamashita, K., Iizuka, S., Taniguchi, N. 1985. A study of zinc distribution in erythrocytes of normal humans.Blut 50:113–116
Peck, E.J., Ray, W.J. 1971. Metal complexes of phosphoglucomutase in vivo. Alterations induced by insulin.J. Biol. Chem. 246:1160–1167
Pfeiffer, D.R., Lardy, H.A. 1976. Ionophore A23187: The effect of H+ concentration on complex formation with divalent and monovalent cations and the demonstration of K+ transport in mitochondria mediated by A23187.Biochemistry 15:935–943
Rifkind, J.M., Heim, J.M. 1977. Interaction of zinc with hemoglobin: Binding of zinc and the oxygen affinity.Biochemistry 16:4438–4443
Scarpa, A., Brinley, F.J., Dubyak, G. 1978. Antipyrylazo III, a “middle range” Ca2+ metallochromic indicator.Biochemistry 17:1378–1386
Simons, T.J.B. 1987. Optical probes for Zn2+ ions.J. Physiol. 394:6P
Simons, T.J.B. 1990. An approach to the estimation of free zinc in human red blood cells.J. Physiol. 423:47P
Smith, R.M., Martell, A.E. 1989. Critical Stability Constants: Second Supplement. Vol. 6. Plenum, New York
Tupper, R., Watts, R.W.E., Wormall, A. 1952. Some observations on the zinc in carbonic anhydrase.Biochem. J. 50:429–432
Van Wouwe, J.P., Veldhuizen, M., DeGoeji, J.J.M., Van den Hamer, C.J.A. 1990. In vitro exchangeable erythrocytic zinc.Biol. Trace Element Res. 25:57–69
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Simons, T.J.B. Intracellular free zinc and zinc buffering in human red blood cells. J. Membrain Biol. 123, 63–71 (1991). https://doi.org/10.1007/BF01993964
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DOI: https://doi.org/10.1007/BF01993964