Acetyl-Coenzyme-A Carboxylase from Rat Liver.

The subunit structure of rat liver acetyl-coenzyme-A carboxylase has been studied by polycrylamide gel electrophoresis in the presence of dodecylsulfate. A number of individual preparations of the enzyme purified by the same procedures exhibited three different types of electrophoretic patterns as follows: first, a single slow-moving protein band (M_r 230 000); secondly two adjacent fast-moving protein bands (M_r 124 000 and 118 000); finally, all three protein bands. With the use of the [¹⁴C]biotin-labelled enzyme, the biotinyl prosthetic group was shown to be associated with the polypeptide of 230 000 M_r as well as with that of 124 000 M_r, but not with the polypeptide of 118 000 M_r. Studies were next made with the labelled enzyme to examine the possibility that the two light polypeptides might have been formed by proteolytic modification of the heavy polypeptide during the procedures used for the purification of the enzyme. Treatment of the enzyme with trypsin or chymotrypsin resulted in cleavage of the heavy polypeptide into two nonidentical polypeptides with molecular weights of approximately 120 000. Incubation of the enzyme with proteases derived from rat liver converted the heavy polypeptide into lighter polypeptides of 80 000-130 000 M_r. Acetyl-CoA carboxylase isolated from crude rat liver extracts by means of immunoprecipitation with specific antibody invariably showed only the heavy polypeptide. The biotin content of the enzyme was found to be 1 mil per 237 000 g protein. These results indicate that rat liver acytyl-CoA carboxylase, unlike bacterial and plant biotin enzymes, has only one kind of subunit, which has a molecular weight of 230 000 and contains one molecule of biotin. Thus, the mammalian enzyme exhibits a highly integrated subunit structure. [ABSTRACT FROM AUTHOR]