The structure and organization of dopamine-beta-hydroxylase in the chromaffin granule membrane.

Chromaffin granules have been purified from bovine adrenal medullae. The granule membranes have been cross-linked with the disulphide-bridged bifunctional imido ester, dimethyl-3,3'-dithiobissuccinimidylpropionate hydrochloride. Analysis of the cross-linked proteins by electrophoresis on agarose/acrylamide gels revealed components of M(r) 300 000 and 150 000. Further analysis of samples by electrophoresis in a second dimension containing a reducing agent revealed the monomeric species from which the cross-linked polypeptides were formed. The major component in the second dimension exhibited a molecular weight of approx. 80 000 and could be identified with dopamine-beta-hydroxylase (3,4-dihydroxyphenylethylamine ascorbate:oxygen oxidoreductase (beta-hydroxylating), EC 1.14.17.1). It is proposed that dopamine-beta-hydroxylase in the intact granule membrane is arranged as a tetramer consisting of two disulphide-bridged dimers of the 80 000 subunit in close apposition. This structural arrangement of the membrane-bound form of dopamine-beta-hydroxylase is identical with that previously proposed for the soluble, intra-granular form of the enzyme.