Genes for catecholamine biosynthesis: cloning by expression and identification of the cDNA for rat dopamine beta-hydroxylase.

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Abstract
MRNA for dopamine .beta.-hydroxylase [3,4-dihydroxyphenylethylamine, ascorbate:oxygen oxidoreductase (.beta.-hydroxylating), EC 1.14.17.1] was partially purified from poly(A)+ mRNA isolated from a rat pheochromocytoma cell line [PC 12]. Shared antigenic determinants between tyrosine hydroxylase and dopamine .beta.-hydroxylase allowed enriched fractions of dopamine .beta.-hydroxylase mRNA to be obtained by immunoprecipitating translated mRNA products with tyrosine hydroxylase antisera. The enriched dopamine .beta.-hydroxylase mRNA was used to synthesize the corresponding c[complementary]DNA which were then cloned in the Pst I site of pBR322. Recombinant colonies were characterized by an in situ colony immunoassay and hybrid-selected translation. In vitro translation of the mRNA selected from 1 recombinant clone produced a protein of 75,000 daltons that comigrated with authetic dopamine .beta.-hydroxylase. Partial proteolysis of authentic dopamine .beta.-hydroxylase and the protein encoded by the recombinant clone produced identical peptide patterns.