PURIFICATION AND SOME PROPERTIES OF CHOLINE ACETYLTRANSFERASE (EC 2.3.1.6) FROM BOVINE BRAIN

Abstract

Abstract— Choline acetyltransferase (ChAc) has been isolated and highly purified from the caudate nuclei of bovine brains. The procedure involved: (1) making an acetone‐ and chloroform‐insoluble powder from the tissue; (2) treating the powder with aqueous buffer and chromatographing the extractable soluble proteins on an organomercurial‐sepharose column; (3) removing impurities by passage through columns of DEAE‐cellulose and hydroxyapatite; and (4) separation of the heme‐containing protein from ChAc by denaturing the former with a mixture of chloroform and n ‐butanol. The purified ChAc was essentially homogeneous as judged by polyacrylamide gel electrophoresis and exhibited a pH optimum at about pH 7. The partially purified ChAc dissociated into two non‐identical subunits when chromatographed with a dilute buffer on Bio‐gel A. It did not dissociate when a more concentrated buffer was used. The purified ChAc dissociated on the Bio‐gel A even in the presence of a high salt concentration. The dissociation was accompanied by a great loss of enzymatic activity, and we concluded that the presence of other proteins tends to prevent the dissociation of ChAc on gel filtration.