Protein-palmitate derivative

Since the protein-palmitate derivative can t be dissolved in organic solvent during homogenization of lipid to form liposomal membranes, it must be inserted into intact liposomes by detergent dialysis. 

Figure 17.20 An azido-palmitic acid derivative can be added to cells to obtain palmitoylated proteins that contain an azide group able to participate in the Staudinger ligation reaction. Biotinylation of these post-translationally modified sites then can be done in vivo using a biotin-phosphine reagent.

In plant systems, de novo synthesis occurs in the plastid and results mainly in the conversion of acetate to palmitate. All 16 carbon atoms in palmitic acid are derived from acetate— half from the methyl carbon and half from the acyl carbon. Two of the carbon atoms (C-15 and C-16) come directly from acetate, and the other 14 come from acetate via the more reactive malonate. Production of malonate requires the incorporation of an additional carbon atom into the acetyl group. This is supplied as bicarbonate, and this same carbon atom is subsequently lost as carbon dioxide. The acyl groups are attached to co-enzyme A (CoASH) during part of the cycle and to acyl carrier protein (ACPSH) during another part. The abbreviated symbols used for these co-enzymes emphasize the thiol groups (SH) to which the acyl chains are attached. 

Fatty acids have predominantly even numbers of carbon atoms because they are effectively formed from acetyl (C2) units, which are derived from glucose in the presence of various enzymes, coenzymes and carrier proteins. An overall scheme for saturated fatty acid biosynthesis is presented in Fig. 2.13, in which it can be seen that the first step is the formation of acetyl coenzyme A (often abbreviated to acetyl-CoA). One molecule ofacetyl-CoA undergoes addition of CO, to form malonyl-CoA, while the acetyl group on another molecule is transferred to an enzyme (fatty acid synthase). The malonyl unit (C3) is added to the enzyme-bound acetyl unit, which produces a butyryl group following loss of C02, dehydration and reduction. Six further steps of combined malonyl addition, decarboxylation, dehydration and reduction occur to yield palmitate (C16). Higher acids are built from palmitate in a similar... 

Fig. 33.11. Regulation of acetyl Co A carboxylase. This enzyme is regulated allosterically, both positively and negatively, by phosphorylation (circled P) and dephosphorylation, and by diet-induced induction (circled t). It is active in the dephosphorylated state when citrate causes it to polymerize. Dephosphorylation is catalyzed by an insulin-stimulated phosphatase. Low energy levels, via activation of an AMP-dependent protein kinase, cause the enzyme to be phosphorylated and inactivated. The ultimate product of fatty acid synthesis, palmitate, is converted to its CoA derivative palmityl CoA, which inhibits the enzyme. A high-calorie diet increases the rate of transcription of the gene for acetyl CoA carboxylase, whereas a low-calorie diet reduces transcription of this gene.

The hepatic uptake of diet-derived copper occurs via the copper transporter 1 (Ctrl), which transports copper with high affinity in a metal-specific, saturable fashion at the hepatocyte plasma membrane (Lee et al., 2001 Klomp et al., 2002). After uptake copper is bound to metallothionein (MT), a cytosolic, low molecular weight, cystein-rich, metal binding protein. MT I and MT II are ubiquitously expressed in all cell types including hepatocytes, and have a critical role to protect intracellular proteins from copper toxicity (Palmiter, 1998 Kelley and Palmiter, 1996). The copper stored in metallothionein can be donated to other proteins. Specific pathways allow the intracellular trafficking and compartmentaUzation of copper, ensuring adequate cuproprotein synthesis while avoiding cellular toxicity (Fig.21.1). 

On the basis of the recent clarification of the function of AGP in fatty acid synthesis, it seems likely that the AGP esters of these intermediate chain length hydroxy acids and unsaturated acids, rather than the CoA esters, are the true intermediates in long-chain saturated and unsaturated fatty acid synthesis. In fact, recently Wakil and co-workers (Wakil et al, 1964 Pugh et al, 1966) have been able to obtain three protein fractions (designated En, Em, and Eiv) involved in the synthesis of palmitic and 11,12-octadecenoic acids. In the presence of acetyl CoA, malonyl CoA, ACP, and TPNH, Eu and Em produced palmitic acid as the major product, while En plus Eiv produced mainly 11,12-octadecenoic acid. Incubation with only En produced a mixture of Cio, Ci2, and C14 3-hydroxyacyl ACP derivatives. Furthermore, the isolated P-hydroxyacyl ACP derivatives could serve as precursors of... 

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