Transferase malonyl

Acyl carrier protein Acyl transferase Malonyl transferase Co-enzyme A... 

The answer is e. (Murray, pp 230-267. Scriver, pp 2297-2326. Sack, pp 121-138. Wilson, pp 287-320.) The fatty acid synthase complex of mammals is composed of two identical subunits. Each of the subunits is a multienzyme complex of seven enzymes and the acyl carrier protein component. All the components are covalently linked together thus, all the components are on a single polypeptide chain, which functions in the presence of another identical polypeptide chain. Each cycle of fatty acid synthesis employs the acyl carrier protein and six enzymes acetyl transferase, malonyl transferase, p-ketoacyl synthase, p-ketoacyl reductase, dehydratase, and enoyl reductase. When the final fatty acid length is reached (usually C16), thioesterase hydrolyzes the fatty acid off of the synthase complex. 

FIGURE 25.10 Acetyl units are covalently linked to a serine residue at the active site of the acetyl transferase in eukaryotes. A similar reaction links malonyl units to the malonyl transferase. 

Consider the role of the pantothenic acid groups in animal fatty acyl synthase and the size of the pantothenic acid group itself, and estimate a maximal separation between the malonyl transferase and the ketoacyl-ACP synthase active sites. 

Production of Malonyl-CoA for the Fatty Acid Biosynthesis. Acetyl-CoA serves as a substrate in the production of malonyl-CoA. There are several routes by which acetyl-CoA is supplied to die cytoplasm. One route is the transfer of acetyl residues from the mitochondrial matrix across the mitochondrial membrane into the cyto-plasm. This process resembles a fatty acid transport and is likewise effected with the participation of carnitine and the enzyme acetyl-CoA-camitine transferase. Another route is the production of acetyl-CoA from citrate. Citrate is delivered from the mitochondria and undergoes cleavage in the cytoplasm by the action of the enzyme ATP-citrate lyase ... 

Fatty-acid synthase (acyl-CoA malonyl-CoA C-acyltransferase, EC 2.3.1.85) is a multifunctional transferase that also has the capacity to hydrolyze thiolesters. The role of its thiolesterase domain is to terminate the growth of fatty acids by hydrolyzing acyl-CoA intermediates [131]. 

Serine hydroxymethyl transferase catalyzes the decarboxylation reaction of a-amino-a-methylmalonic acid to give (J )-a-aminopropionic acid with retention of configuration [1]. The reaction of methylmalonyl-CoA catalyzed by malonyl-coenzyme A decarboxylase also proceeds with perfect retention of configuration, but the notation of the absolute configuration is reversed in accordance with the CIP-priority rule [2]. Of course, water is a good proton source and, if it comes in contact with these reactants, the product of decarboxylation should be a one-to-one mixture of the two enantiomers. Thus, the stereoselectivity of the reaction indicates that the reaction environment is highly hydro-phobic, so that no free water molecule attacks the intermediate. Even if some water molecules are present in the active site of the enzyme, they are entirely under the control of the enzyme. If this type of reaction can be realized using synthetic substrates, a new method will be developed for the preparation of optically active carboxylic acids that have a chiral center at the a-position. 

The enzymatic reaction was performed at 30 °C for 2 hours in a volume of 1 ml of 250 mM phosphate buffer (pH 6.5) containing 50 mM of KOH, 32 U/ml of the enzyme, and [1- C]-substrate. The product was isolated as the methyl ester. When the (S)-enantiomer was employed as the substrate, C remained completely in the product, as confirmed by C NMR and HRMS. In addition, spin-spin coupling between and was observed in the product, and the frequency of the C-O bond-stretching vibration was down-shifted to 1690 cm" (cf. 1740 cm for C-O). On the contrary, reaction of the (R)-enantiomer resulted in the formation of (R)-monoacid containing C only within natural abundance. These results clearly indicate that the pro-R carboxyl group of malonic acid is ehminated to form (R)-phenylpropionate with inversion of configuration [28]. This is in sharp contrast to the known decarboxylation reaction by malonyl CoA decarboxylase [1] and serine hydroxymethyl transferase [2], which proceeds with retention of configuration. 

Spatially, the enzyme activities are arranged into three different domains. Domain 1 catalyzes the entry of the substrates acetyl CoA and malonyl CoA by [ACPj-S-acetyltransferase [1] and [ACPJ-Smalonyl transferase [2] and subsequent condensation of the two partners by 3-oxoacyl-[ACP] synthase [3]. Domain 2 catalyzes the conversion of the 3-0X0 group to a CH2 group by 3-oxoacyl-[ACP]-reductase [4], 3-hydroxyacyl-[ACP -dehydratase [5], and enoyl-[ACP]-re-... 

Figure 3.8 One complete cycle and the first step in the next cycle of the events during the synthesis of fatty acids. ACP = acyl carrier protein, a complex of six enzymes i.e. acetyl CoA-ACP transacetylase (AT) malonyl CoA-ACP transferase (MT) /3-keto-ACP synthase (KS) /J-ketoacyl-ACP reductase (KR) / - hydroxyacyl-ACP-dehydrase (HD) enoyl-ACP reductase (ER).

One enzyme regulated by AMPK is acetyl-CoA carboxylase, which produces malonyl-CoA, the first intermediate committed to fatty acid synthesis. Malonyl-CoA is a powerful inhibitor of the enzyme carnitine acyl-transferase I, which starts the process of ]3 oxidation by transporting fatty acids into the mitochondrion (see Fig. 17-6). By phosphorylating and inactivating acetyl-CoA carboxylase, AMPK inhibits fatty acid synthesis while relieving the inhibition (by malonyl-CoA) of )3 oxidation (Fig. 23-37). 

Malonyl CoA is an allosteric effector of carnitine acyl transferase. What kind of effector is it, i.e., activator or inhibitor, and what is the logic behind the interaction ... 

The pathway can be divided into two metabolic cycles (Figure 3.4). In the first cycle, acetyl-CoA is carboxylated to malonyl-CoA, which is subsequently reduced and converted into propionyl-CoA via 3-hydroxypropionate as a free intermediate. Propionyl-CoA is carboxylated to methylmalonyl-CoA, which is subsequently converted to succinyl-CoA the latter is then used to activate L-malate by succinyl-CoA L-malate coenzyme A transferase, which forms L-malyl-CoA and succinate. Succinate is oxidized to L-malate via conventional steps. L-Malyl-CoA, the second characteristic intermediate of this cycle, is cleaved by L-malyl-CoA/P-methylmalyl-CoA lyase, thus regenerating the starting molecule acetyl-CoA and releasing gly-oxylate as a first carbon-fixation product [27]. 

Two acyl-CoA amino acid A-acyltransferases have been purified from liver mitochondria of cattle, Rhesus monkeys, and humans. One is a benzoyltransferase CoA that utilizes benzyl-CoA, isovaleryl-CoA, and tiglyl-CoA, but not phenylacetyl CoA, malonyl-CoA, or indolacetyl-CoA. The other is a phenylacetyl transferase that utilizes phenylacetyl-CoA and indolacetyl-CoA but is inactive toward benzoyl-CoA. Neither is specific for glycine, as had been supposed from studies using less defined systems both also utilize asparagine and glutamine, although at lesser rates than glycine. 

We were also able to use FAB mass spectrometry to determine the amino acid sequence around the active site serine in the acyl transference domain of rabbit mammary fatty acid synthase.6 The synthase was labelled in the acyl transferase domain(s) by the formation of O-ester intermediates after incubation with [" " C]-acetyl- or malonyl-CoA (Fig. 2A). The modified protein was then digested with elastase (Fig. 2B), and radioactive material isolated via successive purification steps on Sephadex G-50 and reverse phase HPLC. The isolated peptides were then sequenced by FAB MS. The data summarized in Table II established the sequences of both the acetyl and malonyl hexapeptides to be N-acyl-Ser-leu-Gly-Glu-Val-Ala. 

The animal fatty acid synthase (FAS EC 2.3.1.85) is one of the most complex multifunctional enzymes that have been characterized, as this single polypeptide contains all the catalytic components required for a series of 37 sequential transactions (Smith, 1994). The animal FAS consists of two identical polypeptides of approximately 2500 amino acid residues (MW, ca. 270 kDa), each containing seven catalytic subunits (1) ketoacylsynthase, (2) malonyl/acetyl transferase, (3) dehydrase, (4) enoyl reductase, (5) (3-kcto reductase, (6) acyl carrier protein (ACP), and (7) thioesterase. Although some components of the complex are able to carry out their respective catalytic steps in the monomeric form, only in the FAS dimer do the subunits attain conformations that facilitate coupling of the individual reactions of fatty acid synthesis to occur (Smith et al., 2003). 

The entire sequence is described in exquisite detail by Smith et al. (2003 see Figure 2.5). The first step is the sequential transfer of the primer, usually acetyl-CoA, to the serine residue of the acyl transferase, then to the ACP, and finally to (3-ketoacyl synthase. The chain extender substrate, usually malonyl-CoA, is transferred via the serine residue of the acyl transferase to ACP. Condensation is accomplished by (3-ketoacylsynthase, aided by the energetically-favourable decarboxylation of the malonyl residue,... 

Carnitine acyltransferase 1 is strongly inhibited by malonyl CoA, andmuscle has both acetyl CoA carboxylase, which forms malonyl CoA, and malonyl CoA decarboxylase, which acts to remove malonyl CoA and relieve the inhibition of carnitine acyl transferase. The two enzymes are regulated in opposite directions in response to insulin, which stimulates fatty acid synthesis and reduces /S-oxidation, and glucagon that reduces fatty acid synthesis and increases p-oxidation (Kerner and Hoppel, 2000 Louet et al., 2001 Eaton, 2002). 

The activities involved in yeast fatty acid biosynthesis are covalently linked as separate domains of two multifunctional polypeptides, a and p, encoded by the fas2 and fasl genes, respectively (Fig. 2) [57,58]. The functionalities associated with the 220 kDa a subunit include -ketoacyl synthase activity, -ketoacyl reductase activity, and an AGP domain which bears a phosphopantetheinylated serine. The 208 kDa -subunit has acetyl and malonyl CoA transacylase, palmi-toyl transferase, -hydroxyacyl-enzyme dehydratase, and enoyl acyl-enzyme reductase activities. The two subunits can be readily dissociated, and the individual activities maybe measured [57]. 

The seven activities of animal FASs are encoded as separate domains of a single 250 kDa polypeptide (Fig. 2) [30, 31]. These include a -ketoacyl synthase, malonyl/acetyl transferase, -ketoreductase, dehydratase, enoyl reductase, and an ACP domain with a phosphopantetheinylated serine. In addition to these activities, the animal FAS also includes a thioesterase domain which cleaves the product fatty acid from the enzyme. Proteolytic mapping of the polypeptide and genetic analysis have defined the location of the various domains in the primary sequence [30,31]. 

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