Biochemical and molecular biological characterization of plant acetyl-CoA carboxylases
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Abstract
Acetyl-CoA carboxylase is a biotin-containing enzyme which catalyzes the ATP-dependent carboxylation of acetyl-CoA to form malonyl-CoA. Plants are unique among organisms in that malonyl-CoA is utilized in both de novo fatty acid biosynthesis, and as an intermediate in a number of other plant primary and secondary metabolic pathways. Some of these metabolic pathways operate in different subcellular and cellular compartments. Therefore, plants have a mechanism to generate separate pools of malonyl-CoA. Understanding this mechanism has been the impetus for the research reported in this dissertation;Immunological and structural analyses of a 60 kDa biotinylated polypeptide (60kDBP) from carrot indicate that this polypeptide represents the biotin-containing subunit of one isozyme of acetyl-CoA carboxylase. This acetyl-CoA carboxylase exists as a large molecular weight complex, which dissociates upon column chromatography, with a concomitant loss of catalytic activity. In vivo, at least two non-biotinylated polypeptides associate with the 60kDBP. These data indicate that carrot embryos contain a heteromeric form of acetyl-CoA carboxylase, the subunits of which are dissociable;The distribution of acetyl-CoA carboxylase among tissues of leek leaves was determined. The 210 kDa biotinylated subunit of the homomeric acetyl-CoA carboxylase accumulates predominantly in the epidermis of leek leaves. Two subunits of a plastid-localized, heteromeric acetyl-CoA carboxylase in pea leaves are equally abundant in leek mesophyll and epidermal tissues. Thus, leek leaves contain a homomeric and a heteromeric isozyme of acetyl-CoA carboxylase, the former being restricted almost exclusively to the epidermis. The homomeric acetyl-CoA carboxylase may be involved in the elongation of long-chain fatty acids in the leek epidermis;A partial cDNA for the 220 kDa soybean acetyl-CoA carboxylase isoform has been isolated. The deduced amino acid sequence is highly homologous to the deduced sequences of acetyl-CoA carboxylase genes from plants, animals, and algae. Northern blot analysis suggests that the expression of the soybean gene may be controlled in a tissue-specific manner. The soybean genome contains two to four copies of this gene.