COPURIFICATION, CO-IMMUNOPRECIPITATION, AND COORDINATE EXPRESSION OF ACETYL-COENZYME-A CARBOXYLASE ACTIVITY, BIOTIN CARBOXYLASE, AND BIOTINCARBOXYL CARRIER PROTEIN OF HIGHER-PLANTS

Acetyl-CoA carboxylase (ACCase; EC 6.4.1.2) is a regulatory enzyme of fatty acid synthesis, and in some higher-plant plastids is a multi-sub unit complex consisting of biotin carboxylase (BC), biotin-carboxyl ca rrier protein (BCCP), and carboxyl transferase (CT). We recently descr ibed a Nicotiana tabacum L. (tobacco) cDNA with a deduced amino acid s equence similar to that of prokaryotic BC. We here provide further bio chemical and immunological evidence that this higher-plant polypeptide is an authentic BC component of ACCase. The BC protein co-purified wi th ACCase activity and with BCCP during gel permeation chromatography of Pisum sativum L. (pea) chloroplast proteins. Antibodies to the Rici nus communis L. (castor) BC co-precipitated ACCase activity and BCCP. During castor seed development, ACCase activity and the levels of BC a nd BCCP increased and subsequently decreased in parallel, indicating t heir coordinate regulation. The BC protein comprised about 0.8% of the soluble protein in developing castor seed, and less than 0.05% of the protein in young leaf or root. Polypeptides cross-reacting with antib odies to castor BC were detected in several dicotyledons and in the mo nocotyledons Hemerocallis fulva L. (day lily), Iris L., and Allium cep a L. (onion), but not in the Gramineae species Hordeum vulgare L. (bar ley) and Panicum virgatum L. (switchgrass). The castor endosperm and p ea chloroplast ACCases were not significantly inhibited by long-chain acyl-acyl carrier protein, free fatty acids or acyl carrier protein. T he BC polypeptide was detected throughout Brassica napus L. (rapeseed) embryo development, in contrast to the multifunctional ACCase isoenzy me which was only detected early in development. These results firmly establish the identity of the BC polypeptide in plants and provide ins ight into the structure, regulation and roles of higher-plant ACCases.