Abstract
A low molecular weight, native zinc binding, cytosolic protein (LMZP) has been isolated, purified and characterized from human normal term placenta. Gel filtration of heat treated placental cytosol after sequential acetone precipitation (80% ppt) revealed a major zinc binding protein in the range of low molecular weight. This partially purified zinc binding fraction was further fractionated on DEAE-Sephadex A-25. The zinc was eluted in one of the three peak fractions. Further, the purity of zinc binding protein was confirmed on fast protein liquid chromatography (FPLC). The purified placental LMZP was homogenous on SDS-polyacrylamide gel electrophoresis with a single band. Ultraviolet (UV) spectrum of LMZP showed an absorption maximum at 257 nm which disappeared at pH 2. Molecular weight of LMZP as determined by gel chromatography, SDS-polyacrylamide gel electrophoresis and amino acid analysis was 6 kDa. It was calculated that 1 g atom of zinc was bound to 1 mole of the LMZP. Unlike in classical metallothionein, the amino acid composition of placental LMZP revealed the presence of aromatic amino acids, lower content of cysteine and higher content of histidine, glutamic acid and aspartic acid (10, 9 and 5 residues/mole, respectively).
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Hughes MN: The study of metalloproteins and other metal-containing biological molecules. In: The inorganic chemistry of biological processes. John Wiley Inc., New York, 1972, pp 63–104
Hanzlik RP: Inorganic aspects of biological and organic chemistry. Academic Press, New York, 1976, pp 215–251
Vallee BL, Wacker WEC: Metal protein interactions. In: H. Neurath (ed). Academic Press, New York, 1970, pp 25–26
Nomiyama K, Nomiyama H: High-performance liquid chromatographic determination of tissue metallothionein in monkeys chronically exposed to cadmium. J Chromatogr 228: 285–289, 1982
Giroux E, Lachmann PJ: Kidney and liver metallothioneins in rats after administration of an organic compound. J Biol Chem 259: 3658–3662, 1984
Anderson RA, Eriksen KDH, Bakke T: Evidence of presence of a low molecular weight, non-metallothionein-like metal-binding protein in the marine Gastropodnassarius reticulatus L. Comp Biochem Physiol 94B: 285–291, 1989
Leber AP, Miya TS: A mechanism for cadmium-and zinc-induced tolerance to cadmium toxicity: Involvement of metallothionein. Toxicol Appl Pharmacol 37: 403–414, 1976
Probst GS, Bousquet WF, Miya TS: Correlation of hepatic metallothionein concentrations with acute cadmium toxicity in the mouse. Toxicol Appl Pharmacol 39: 61–69, 1977
Danielson B, Hanssoun E, Dencker L: Placental transport of chromium (Cr) and its effect on cartilage formation. Teratology 26: 47A-50A, 1982
Bremner I, Williams RB, Young BW: Distribution of copper and zinc in the liver of the developing sheep fetus. Br J Nutr 38: 87–92, 1977
Riordan JR, Richards V: Human fetal liver contains both zinc and copperrich forms of metallothionein. J Biol Chem 255: 5380–5384, 1980
Kern SR, Smith HA, Fontaine D, Bryan SE: Partitioning of zinc and copper in fetal liver subfractions: Appearance of metallothionein-like proteins during development. Toxicol Appl Pharmacol 59: 346–354, 1981
Lucis OJ, Lucis R, Shaikh ZA: Cadmium and zinc in pregnancy and lactation. Arch Environ Health 25: 14–22, 1972
Waalkes MP, Poisner AM, Wood GW, Klaassen CD: Metallothionein-like proteins in human placenta and fetal membranes. Toxicol Appl Pharmacol 74: 179–184, 1984
Arizono K, Otas S, Ariyoshi T: Purification of metallothionein-like protein in rat placenta. Bull Environ Contam Toxicol 27: 671–677, 1981
Wong K, Klaassen CD: Isolation and characterization of metallothionein which is highly concentrated in newborn rat liver. J Biol Chem 254: 12399–12403, 1979
Waalkes MP, Bell JU: Isolation and partial characterization of native metallothionein in the fetal rabbit liver. Life Sci 27: 585–593, 1980
Vig PJS, Paliwal VK, Nath R: A comparative study of direct current plasma, atomic emission spectrometry and atomic absorption spectrophotometer for biological monitoring of trace metals. In: Dhillon HK and Hu MH (eds) Biological monitoring of exposure to chemicals and metals. John Wiley and Sons, Inc, New York, 1991, pp 163–171
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ: Protein measurement with the Folin phenol reagent. J Biol Chem 193: 265–175, 1951
Schales O, Schales SS: A simple and accurate method for the determination of chloride in biological fluids. J Biol Chem 140: 879–884, 1941
Laemmli UK: Cleavage of structural proteins during the assembly of the head of bacterophage T4. Nature 227: 680–685, 1970
Du Val G, Fowler BA: Preliminary purification and characterization studies of a low molecular weight, high affinity cytosolic lead-binding protein in rat brain. Biochem Biophys Res Commun 159: 177–184, 1989
Ellman GL: Tissue sulfhydryl groups. Arch Biochem Biophys 82: 70–77, 1959
Law AYC, Stillman MJ: The effect of pH on Cd+2 binding to rat liver metallothionein. Biochem Biophys Res Commun 94: 138–143, 1980
Paliwal VK, Kohli KK, Sharma M, Nath R: Purification and characterization of metallothionein from liver of cadmium exposed rhesus monkeys (Macaca mulatta). Mol Cell Biochem 71: 139–144, 1986
Pantolino MW, McDonnel PJ, Valentine JS: Reversible loss of metal ions from the zinc binding site of copper-zinc superoxide dismutase. The low pH transition. J Am Chem Soc 101: 6454–6456, 1979
Waalkes MP, Chernoff SB, Klaassen CD: Cadmium-binding proteins of rat testes: Characterization of a low-molecular-mass protein that lacks identity with metallothionein. Biochem J 220: 811–818, 1984
Sarkar B: Metal protein interactions. Prog Food Nutr Sci 11: 363–400, 1987
Lipscomb WN, Hartsuck JA, Quiocho FA, Reeke GN: The structure of carboxypeptidase A. IX. The X-ray diffraction results in the light of the chemical sequence. Proc Natl Acad Sci 64: 28–35, 1969
Richardson JS, Thomas KA, Rubin BH, Richardson DC: Crystal structure of bovine Cu, Zn superoxide dismutase at 3Å resolution. Proc Natl Acad Sci 72: 1349–1353, 1975
Chen RW, Ganther HE: Some properties of a unique cadmium-binding moiety in the soluble fraction of rat testes. Environ Physiol Biochem 5: 235–238, 1975
Charles-Shannon VL, Sasser LB, Burbank BK, Kelman BJ: The influence of zinc on the ontogeny of hepatic metallothionein in the fetal rat. Proc Soc Exp Biol Med 168: 56–61, 1981
Greeley S, Johnson T, Schafer D, Johnson PE: Gestational alcoholism and fetal zinc accration in Long-Evans rats. J Am Coll Nutr 9: 265–271, 1990
Seal CJ, Heaton FW: Zinc transfer among proteins in rat duodenum mucosa. Ann Nutr Metab 31: 55–60, 1987
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Honey, S., Dhall, G.I. & Nath, R. Purification and characterization of a low molecular weight zinc binding protein from human placenta. Mol Cell Biochem 136, 77–83 (1994). https://doi.org/10.1007/BF00931608
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DOI: https://doi.org/10.1007/BF00931608