Role of non-catalytic subunits in biotin-dependent carboxylases

AcCCase play an important role in the production of malonyl-CoA for fatty acid synthase (FASand polyketide synthase (PKS) in bacteria, plants and fungi. Many of the polyketides produced with malonyl-CoA are pharmaceutically important. Fatty acids function as building blocks of membranes and are also used in the biosynthesis of wide array signaling molecules and triglycerides. AcCCase, specifically ACCase, was shown to be essential for the normal growth of the Saccharomyces, E. coli and B. subtilis. ACCase catalyzes the first committed reaction of de novo fatty acid biosynthesis which is catalyzed by FAS. The malonyl CoA produced by ACCase is first transformed to malonyl-ACP via the action of malonyl-CoA-ACP transacylase. The initial fatty acid synthesis reaction is the condensation of malonyl ACP (the acyl-ACP extender substrate) with an acyl-CoA starter substrate (usually acetyl-CoA), via the action of ketoacyl-ACP synthase to generate a β-ketoacyl-ACP intermediate. Via a multi-step cycle, this β-ketoacyl-ACP is reduced, dehydrated and subsequently reduced to produce a saturated acyl ACP that can serve as the extender substrate for subsequent rounds of condensation and elongation with another malonyl-ACP unit. In most organisms the products of the fatty acid synthesis cycle are fatty acids of 16 or 18 carbons in length. These fatty acids play important roles in cellular membranes, organellar membranes, as energy storage compounds, and as signaling molecules. The involvement of biotin-dependent carboxylases in metabolic pathways has made them a new target for the development of antibiotics , and recognized as potential targets for cancer, obesity and herbicides . Currently, solved structures of many holoenzymes and subunits/domains of biotin-dependent carboxylases exhibit a diverse enzyme architecture, even though they share a significant amount of sequence similarity, especially at the BC and CT components. This is an evidence of diversity existing in nature. Consequently, understanding the structure and functional relationship between subunits of each biotin-dependent carboxylases has become the primary interest of the scientific community. This study is focused on characterization of non-catalytic subunits of biotin dependent carboxylases using co-purification of recombinant proteins expressed in the bacterial expression system and other relevant enzyme kinetic assays