Serine transacetylase

The biosynthesis of L-cysteine entails the sulfhydryl transfer to an activated form of serine. This pathway to L-cysteine has been most thoroughly studied in E. coli. In the first step an acetyl group is transferred from acetyl-CoA to serine to yield (9-acetylserine (fig. 21.8a). The reaction is catalyzed by serine transacetylase. The formation of cysteine itself is catalyzed by O-acetylserine sulfhydrylase. 

Fig. 2. Summary of the free and bound pathways of sulfate assimilation in plants. Some related reactions and points of entry of several forms of inorganic sulfur are also shown. The reaction sequence catalyzed by (1) ATP sulfurylase, (2) APS sulfotransferase, (3) thiosulfonate reductase, and (4) cysteine synthase constitutes the bound sulfate assimilation pathway. The synthesis of OAS is catalyzed by (5) serine transacetylase. The reaction sequence (I), (6)-(9)or (1), (2), (10), (8), (9) constitutes the free pathway reactims (7) and (10) are nonenzymatic, (6) is catalyzed by APS sulfotransferase, (8) by sulfite reductase, and (9) by cysteine synthase. APS and PAPS are interrelated via (11) APS kinase and (12) NDP phophohydrolase. APS can be hydrolyzed via (13) APS sulfohydrolase or (14) APS cyclase.

The enzyme is specific for OAS. Since plants contain the enzyme serine transacetylase (Smith and Thompson, 1971) and tobacco cultures synthesize OAS from serine (Smith, 1977), it is presumed that OAS serves as the physiological sulfide acceptor (Giovanelli et al., this volume. Chapter 12). 

A possible minor route of serine degradation could be via the conversion of serine to O-acetyl serine by serine transacetylase which has been partially purified from kidney bean seedlings by Smith and Thompson (1971). O-acetyl... 

Figure 12.17 Important targets for the generation of an L-cysteine production strain. Mutant proteins are marked with an asterisk. Exp. cysteine export proteins. SerA 3-phosphoglycerate dehydrogenase. CysE serine transacetylase. CysB transcriptional activator of the cystein regulon. CysK O-acetyl-L-serine (thiol)-lyase A. CysM O-acetyl-L-serine (thiol)-lyase B.

The second important feedback mechanism which has to be eliminated regulates the penultimate reaction in L-cysteine biosynthesis. The L-serine transacetylase CysE is normally inhibited effectively by the end-product L-cysteine (Kj 1 pM). Also for this enzyme various muteins were generated showing a very efficient decoupling of feedback regulation by the end-product. Most powerful is the mutant allel named cysE23 encoding a protein with a K value of 2.3 mM. ... 

An alternative issue for avoidance of feedback inhibition by L-cysteine is the use of heterologous L-serine transacetylases. Very promising proteins can be found in plants which are summarized in the review by Wada and Takagi. ... 

The effect of different amino acids supplements on the synthesis of PHB by recombinant E. coli was evaluated by Mahishi and Rawal. The study revealed that when the basal medium is supplemented with amino acids, except glycine and valine, all other amino acid supplements enhanced PHB accumulation in recombinant E. coli harboring PHB synthesizing genes from S. aureqfaciens. Cysteine, isoleucine, or methionine supplementation increased PHB accumulation by 60, 45, and 61%, respectively. Amino acid biosynthetic enzyme activities in several pathways are repressed by end produa supplementation. End product inhibition in the cysteine biosynthetic pathway controls the carbon flow due to sensitivity of serine transacetylase to cysteine. Hence, supplementation of cysteine favors a change in carbon flux that eliminates the requirement of acetyl-CoA for serine transacetylation which in turn provides more carbon source and acetyl-CoA for PHB synthesis. Degradation of methionine and isoleucine yields succinyl CoA, an intermediate of tricarboxylic acid cycle and allows more acetyl-CoA to enter the PHB biosynthetic pathway. 

The amino acid cysteine is essential for wool growth in sheep, which need a source of methionine to synthesise it. Certain bacteria have the ability to synthesise cysteine, with the pathway involving the action of two enzymes, serine transacetylase and O-acetylserine sulphydralase. The genes coding for these enzymes have been successfully introduced into sheep, which then express the appropriate pathways but, so far, only in inappropriate tissues. 

Fig. 3. Regulation of the bound pathway for the assimilation of sulfate into cysteine and associated processes. Carrier refers to an endogenous thiol of uncertain identity in higher plants. Enzymes associated with the sulfate assimilation pathway and the synthesis of O-acetylseiine are (1) high-ailinity sulfate uptake mechanism, (2) ATP-sulfurylase, (3) adenosine S -phosphosulfate (APS) sulfotransferase, (4) organic thiosulfate reductase, (5) cysteine synthase, and (6) serine transacetylase. Cysteine sulfhydrase (7), an enzyme of cysteine catabolism, and nitrate reductase (8), the first enzyme of the nitrate assimilation pathway, are also shown. Inhibitory control of the pathways is shown by discontinuous lines (----) and enhancement by continuous lines (------).

O-Acetylserine is synthesized from serine and acetyl-coenzyme A in a reaction catalyzed by serine transacetylase. Brunold and Suter (1982) reported that about 35% of the activity present in spinach leaves is associated with chloroplasts. Since no activity was associated with mitochondria or peroxisomes the remaining 65% is presumably associated with the cytosol. Brunold and Suter (1982) also reported that the enzyme is almost completely inactive in the presence of 1 mM L-cysteine, confirming the earlier obrervation of Smith and Thompson (1971). Some further details of cysteine syntha.se, mostly concerning amino acid composition and the presence of multiple forms, have been reported since the previous review in this series (Diessner and Schmidt, 1981 Ikegami et al, 1987 Murakoshi et al, 1985,1986). 

The internal regulation of the sulfate assimilation pathway and its coordination with the nitrate assimilation pathway are summarized in Fig. 3. It shows that cysteine is a negative effector of serine transacetylase and that it also controls the level of APS sulfotransferase. The inhibitory effects of HjS on the level of APS sulfotransferase are probably mediated via cysteine, though HjS itself at high concentrations inhibits cysteine synthase. 

Fig. 6. Subcellular location of the enzymes of cysteine and methionine synthesis in photosynthetic cells in C3 plants. The scheme incorporates the proposals of Wallsgrove et al. (1983) for the synthesis of phosphohomoserine and methionine and current understanding of the location of the enzymes of the sulfate assimilation pathway and of serine transacetylase. Abbreviations OAS, 0-acetylserine PHS, phosphohomoserine THF, tetrahydrofolate.

PDH is a multi-enzyme complex consisting of three separate enzyme units pyruvate decarboxylase, transacetylase and dihydrolipoyl dehydrogenase. Serine residues within the decarboxylase subunit are the target for a kinase which causes inhibition of the PDH the inhibition can be rescued by a phosphatase. The PDH kinase (PDH-K) is itself activated, and the phosphatase reciprocally inhibited, by NADH and acetyl-CoA. Figure 3.12(a and b) show the role and control of PDH. 

Control of pymvate dehydrogenase activity is via covalent modification a specific kinase causes inactivation of the PDH by phosphorylation of three serine residues located in the pyruvate decarboxylase/dehydrogenase component whilst a phosphatase activates PDH by removing the phosphates. The kinase and phosphatase enzymes are non-covalently associated with the transacetylase unit of the complex. Here again we have an example of simultaneous but opposite control of enzyme activity, that is, reciprocal regulation. 

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