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It has been previously shown that complexation of Cu2+ is essential for effective uptake of Cu2+ by brain tissues and that 67Cu complexed to His is taken up by a high affinity and a low affinity saturable process (Hartter, D. E., and Barnea, A. (1988) J. Biol. Chem. 263, 799-805). Using rat hypothalamic tissue slices, we defined the ligand specificity for these two uptake processes. The effectiveness of stereoisomers or methyl (Me) derivatives of His in facilitating 67Cu uptake by the high affinity process was in this decreasing order: L-His = D-His = Me-3-N-His greater than Me-ester-His greater than Me-alpha-N-His greater than or equal to Me-1-N-His. By the low affinity process it was: L-His = D-His = Me-3-N-His = Me-ester-His = Me-alpha-N-His greater than Me-1-N-His. When facilitation of 67Cu uptake by 14 different amino acids was evaluated using copper:ligand (Cu:L) ratios of 1:2,000 (high affinity process) or 1:2 (low affinity process), His stood out as the most effective. However, when [Cu2+] was 0.1 microM and the Cu:L ratio was increased from 1:2,000 to 1:20,000, Ala, Gly, Lys, Ser, or Thr was each as effective as His; when [Cu2+] was 10 microM and the Cu:L ratio was increased from 1:2 to 1:2,000, Gln, Glu, Gly, Lys, or Ser was each superior to His in facilitating 67Cu uptake. Moreover, by comparison to 67Cu uptake at a Cu:L ratio of 1:2, increasing the ratio attenuated (His) or enhanced (Gln, Glu, Gly, Lys, Ser) 67Cu uptake. These results indicate that 1) coordination of Cu2+ with the 1-N-imidazole and the alpha-amino (but not with the carboxyl) is essential for His facilitation of 67Cu uptake, and 2) the amino acid specificity for uptake of complexed Cu2+ is a function of both [Cu2+] and the molar ratio of copper to amino acid. These results are consistent with coordination of Cu2+ with at least three nitrogens being a primary factor facilitating copper uptake by brain tissue. |
