Abstract
This chapter focuses on four enzymes, which include citrate synthase, ATP citrate lyase, citrate lyase, and isocitrate lyase. All of these enzymes catalyze reversible aldol condensations. The first three enzymes have citrate as their substrate. They act on citrate to cleave it into oxalacetate and acetate or acetyl-CoA. Despite these and other gross similarities, the citrate-cleaving enzymes are separate and distinct from one another. When they occur in the same cell, precise compartmentation secures their separate function. Thus in mammalian cells, citrate synthase is mitochondrial, while ATP citrate lyase is cytoplasmic, in localization. Isocitrate, a structural isomer of citrate, undergoes, like citrate, an aldolic cleavage into a four-carbon (succinate) and a two-carbon (glyoxylate) fragment in the presence of isocitrate lyase. A straightforward chemical assay of citrate synthase depends on the colorimetric determination of the citrate, which is formed in the reaction. Citrate synthases have been isolated from pig heart, pigeon breast muscle, moth flight muscle, beef liver and beef heart, yeast, lemon, cauliflower and bean seedlings, rat liver, and several bacteria. ATP citrate lyase is found in pigeon liver, beef brain and pig heart, certain bacteria, and several tissues of rat, rabbit, and chicken. For assay of ATP citrate lyase, two methods are commonly used. The first depends on trapping the acetyl-CoA formed in citrate cleavage as acetyl hydroxamate, and measuring the latter by the color produced with ferric chloride. A second, more refined ,assay monitors oxalacetate, the other citrate cleavage product, by its reaction with NADH in the presence of malate dehydrogenase.
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