Citric acid enzymatic synthesis

Stem JR, Ochoa S (1951) Enzymatic synthesis of citric acid 1. Synthesis with soluble enzymes. JBiol Chem 191 161-172... 

Another observation on oxalate formation is that other a-keto acids, such as oxalosuccinic acid (74) and a-ketoglutaric acid (106) do not seem to yield oxalate directly but indirectly (123). This appears to be due to the fact that only oxaloacetic acid can function as an acetate donor. In this connection the intervention of Coenzyme A may be considered, since it is reported to function in the acetylation of sulfanilamide and choline (73) and recently was shown to take part in the enzymatic synthesis of citric acid. This concept may be illustrated as follows ... 

The first problem was solved 133) through synthesis (followed by enzymatic degradation) of tritiated citric acid stereospecifically labeled in the pro-R branch, as indicated in Fig. 62. The starting material for this synthesis is the naturally occurring 5-dehydro-... 

Net Synthesis of a-Ketoglutarate a-Ketoglutarate plays a central role in the biosynthesis of several amino acids. Write a sequence of enzymatic reactions that could result in the net synthesis of a-ketoglutarate from pyruvate. Your proposed sequence must not involve the net consumption of other citric acid cycle intermediates. Write an equation for the overall reaction and identify the source of each reactant. 

It is well known that in the oxidation of pyruvate an active 2-carbon intermediate, known as active acetate, is formed (Gurin and Crandall, 1948 Bloch, 1948 Lehninger, 1950). A suggestion that pantothenic acid is involved in the conversion of acetate to citrate in yeast came from Novelli and Lipmann s (1950) experiments showing a correlation between the ability of the yeast to oxidize acetate or ethanol and the coenzyme A content of the cells. The formation of citrate from acetate and oxaloacetate in cell-free extracts from pigeon liver or yeast was also shown to require coenzyme A (Novelli and Lipmann, 1950 Stern and Ochoa, 1949, 1951). Further study of these enzyme systems led to the isolation of a crystalline condensing enzyme from pig heart (Ochoa et al., 1951) and to a formulation of the mechanism of the enzymatic synthesis of citric acid from... 

Reaction (a) is the sum of at least two reactions. It is catalyzed by enzyme fractions A and B and also requires diphosphothiamine. Reaction (a) can be coupled with either reaction (b), in which acetylphosphate is formed in the presence of transacetylase, or with reaction (c), which requires the condensing enzyme. In animal tissues, which contain no transacetylase, reaction (a) is coupled with reaction (c). Further discussion of the enzymatic mechanisms involved in the synthesis of citric acid can be found in reviews by Krebs (1949), Ochoa (1951), and Barker (1951). 

Korkes, S., del Campillo, A., Gunsalus, I. C. and Ochoa, S. (1951) Enzymatic synthesis of citric acid. IV. Pyruvate as acetyl donor. J. Biol. Chem. 193, 721-735. 

There is a number of similar procedures starting fntm olefinic diols which by reaction with dinitrogen tetraoxide give citric acid. Sargsyan et al. [11] presented a short account of all known until 1989 synthetic preparations of citric acid. Their paper is based mainly on the patent literature and shows that with an exception of old classical methods, most of other ways to obtain citric acid is characterized by relatively low yield. Evidently, in the context of the Krebs tricarboxylic acid cycle, there is a large number of investigations dealing with enzymatic synthesis of citric acid by condensation of acetate and oxalacetate [12-20]. 

Ochoa S, Stern JR, Schneider MC (1949) Enzymatic synthesis of citric acid. II. Crystalline condensing enzyme. J Biol Chem 179 491-492... 

---