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ARS Home » Midwest Area » West Lafayette, Indiana » Crop Production and Pest Control Research » Research » Publications at this Location » Publication #94041


item Nielsen, Niels

Submitted to: Plant Physiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/16/1998
Publication Date: N/A
Citation: N/A

Interpretive Summary: Because oil is an important product derived from soybean, understanding the mechanisms that regulate its synthesis is important. The chloroplast enzyme called acetyl CoA carboxylase is responsible for a key reaction during formation of the fatty acids used for oil biosynthesis. This communication describes the four components that comprise the soybean enzyme. They are called biotin carboxylase (BC), biotin carboxyl carrier protein (BCCP), alpha-carboxytransferase (alpha-CT) and beta-carboxytransferase (beta-CT). We show that BC, BCCP and alpha-CT are each produced at the direction of several different DNAs. This will complicate efforts to manipulate the activity of the enzyme. We also demonstrate that BC/BCCP and alpha-CT/beta-CT each form extremely large complexes that when added together restore enzyme activity. The complicated structures that are assumed by components of acetyl CoA carboxylase could reflect a mechanism that regulates the activity of the enzyme. The information in this communication is directed toward specialists doing research in this area, and may be useful during efforts to improve the oil content of soybean.

Technical Abstract: A multisubunit form of acetyl CoA carboxylase (ACCase) from soybean was characterized. The enzyme catalyzes the formation of malonyl CoA from acetyl CoA. The four known components that comprise plastid ACCase are biotin carboxylase (BC), biotin carboxyl carrier protein (BCCP), and the alpha- and beta-subunits of carboxytransferase (alpha- and beta-CT). At least three different cDNAs were isolated from germinating soybean seeds that encode BC, two that encode BCCP, and four that encode alpha-CT. Whereas BC, BCCP and alpha-CT are products of nuclear genes, DNA that encodes soybean beta-CT is located in chloroplasts. Translation products from cDNAs for BC, BCCP and alpha-CT were imported into isolated pea chloroplasts and became integrated into ACCase. Edman microsequence analysis of the subunits after import permitted identification of the amino terminal sequence of the mature protein. Antibodies specific for each of the chloroplast ACCase subunits were generated, and permitted components of ACCase to be followed during fractionation of the chloroplast stroma. In the presence of 0.5 M KCl, a complex that contained BC plus BCCP emerged from Sephacryl 400 with an apparent molecular weight greater than 800 kDa. A second complex, which contained alpha- and beta-CT, was also recovered, and had a molecular weight greater than 600 kDa. By mixing the two complexes at appropriate ratios, ACCase enzymatic activity was restored. Even higher ACCase activities were recovered by mixing complexes from pea and soybean. The results suggest that the active form of ACCase is an extremely high molecular weight complex.