Catecholamine Binding Macromolecule in Soluble Fraction of Rat Brain

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Abstract
A macromolecule having a high binding affinity to norepinephrine was isolated from the soluble fraction of rat brain, and the partially purified macromolecule was prepared by extraction of the brain with isotonic sucrose containing 3.3 mM CaCl2, ultracentrifugation, 35-50% (NH4)2SO4 precipitation and DEAE-cellulose chromatography. DEAE-cellulose chromatography revealed that the macromolecule in the soluble fraction was separated in at least 4 fractions, 2 of which had both norepinephrine and cyclic(c)AMP binding activity. A Scatchard plot indicated that a partially purified macromolecule showed 2 apparent binding affinity constants of 4.13 .times. 10-7 M and 3.70 .times. 10-6 M for norepinephrine. The effect of chemical or physical treatments on specific binding of the macromolecule to norepinephrine was studied in comparison with cAMP binding to the macromolecule. Trypsin treatment caused partial loss of norepinephrine binding activity, while cAMP binding activity was completely lost. Heat treatment resulted in a complete disappearance of cAMP binding activity, while norepinephrine binding activity was only slightly decreased. Dibenamine treatment caused a marked decrease of norepinephrine and cAMP binding activity. Dichloroisoproterenol had no effect on the binding activities. cAMP clearly inhibited norepinephrine binding to the macromolecule, while norepinephrine inhibited cAMP binding only slightly. Norepinephrine binding to the macromolecule was essentially dependent on its catechol structure. The macromolecule was discussed on the basis of the findings.

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