Yeast acetoacetate activation

An optically active reagent 32 for the left hand fragment can be made by asymmetric reduction of ethyl acetoacetate with ordinary baker s yeast (see chapter 29), protection of the OH group as a mixed acetal 31, reduction of the ester and conversion of the new OH group into an iodide 32. 

A mild and inexpensive way to reduce aldehydes or ketones uses fermenting Baker s yeast. This is a whole-cell system that contains oxidoreductase enzymes and cofactors that reduce the substrate. The ketonic carbonyl groups of (3-keto-esters and cyclic ketones are reduced with high selectivity using Baker s yeast. Typical in this regard is the reduction of ethyl acetoacetate, which gives ethyl 3-hydroxybutyrate as predominantly the (5)-stereoisomer (7.101). Similarly, the ketone 114 gave the optically active 3-hydroxyproline derivative 115 (7.102). 

A variety of thiokinases probably exist, but only a few of them have been identified. Acetic acid and butyryl thiokinase have been purified from a variety of sources, including yeast, liver, and muscle. These two enzymes differ in their specificity for the substrate. Acetic thiokinase catalyzes only the oxidation of propionic, acetic, and acrylic acids, but butyryl thiokinase activates fatty acids of chain lengths ranging from 4-to 12-carbon units. A third thiokinase was also discovered. It acts on fatty acid chains with 5- to 22-carbon units and is found in the microsomes. This intracellular distribution is in striking contrast with the cellular location of all other enzymes involved in fatty acid oxidation, which are all in mitochondria. The palmityl enzyme, which is active in the presence of ATP and CoA, becomes inactive when incubated in the absence of CoA therefore, it has been proposed that the active form of the enzyme involves the formation of an enzyme-CoA complex. The heart, the skeletal muscle, and the kidney also contain a thiokinase that specifically activates acetoacetic acid. Acetoacetic acid thiokinase is absent in liver this observation is significant in the pathogenesis of ketosis. 

As the study of CoA developed, it became apparent that the coenzyme was involved in reactions whereby acetate was activated by ATP and subsequently transferred to various acetyl acceptors. In pigeon liver extracts it was shown that acetate could be activated by ATP in the presence of CoA to acetylate sulfanilamide, PABA, histamine, glucosamine, to synthesize acetoacetic acid and citrate. Acetyl phosphate, which has been demonstrated to be a product of pyruvate metabolism in several bacteria and could theoretically be considered to be an intermediate in these reactions, was found to be unable to replace acetate and ATP in animal tissues. Eventually it was shown that there is present in certain bacteria an enzyme, phosphotransacetylase, which could convert acetyl phosphate to a reactive product which was thought to be acetyl-CoA.i 194 isolation of acetyl-CoA from yeast extract by Lynen and Reichert confirmed the idea that acetyl-CA is the reactive 2-carbon unit in these reactions. Stadtman has demonstrated that acetyl-CoA is indeed the product of the action of phosphotransacetylase. Lipmann has recently... 

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