Citrate synthetase

Among fluorinated secondary metabolites, the following examples can be examined (Figure 4.1). Fluorocitrate comes from the in vivo conversion of fluoroacetate and is responsible for the toxicity of this latter compound (cf. Chapter 7). Condensation of fluoroacetyl-CoA with oxaloacetate is catalyzed by citrate synthetase (this enzyme usually incorporates Ac-CoA in the cycle of the citric acid). " ... 

Scheme 18. Biosynthesis of (2R, 3 )-fluorocitric acid with citrate synthetase...

Acetic acid measurement involves conversion of acetate to acetyl-CoA by acetyl-CoA synthetase with the consumption of ATP (Boehringer Mannheim, 1986). Acetyl-CoA reacts with oxaloacetate and water in the presence of citrate synthetase to form citrate and CoA. Oxaloacetate for this reaction is obtained from malate by the action of malate dehydrogenase with concomitant conversion of NADH from NAD+. NADH is spectrophotometrically measured and correlated to acetic acid concentration. 

This is not true for all citrate synthetases enzymes from some sources lead to addition to the Re face >36). This finding that enzymes from different sources promote one and the same reaction in stereochemically distinct fashion is by no means unique. 

The ability to analyze CHDTC02H stereochemically was put to use in the elucidation 156,157) of the stereochemistry of the citric acid condensation to which we have already alluded. Before presenting one of the determinations of the stereochemistry of the citrate synthetase reaction, however, we must develop that of the reverse process, the citrate lyase reaction156). The essential features are shown in Fig. 69, bottom. The key to this determination is the synthesis of citric acid stereo-... 

With the stereochemistry of the citrate lyase reaction determined, that of the Si citrate synthetase (the common enzyme) was established as shown in Fig. 70. Condensation of (J )-acetic-d, t acid (configuration known by synthesis) with oxalo-acetate gives what turns out to be mainly (2S,3/ )-citric-2-d,2-/ acid (112).41 When this acid is then cleaved with citrate lyase, the major product is (/ )-acetic-d, t acid, as established by the malate synthetase/fumarase diagnosis. It follows that both the Si-citrate synthetase and citrate lyase reactions must involve the same stereochemical course. Since that of the lyase reaction is inversion (vide supra), that of the Si synthetase reaction must be inversion also. And since the overall stereochemical result shown in Fig. 70 is not dependent on the magnitude of the... 

The effect of nonfatal injuries such as a 2-hour period of bilateral hind-limb ischemia or a full-thickness scald of 20% of skin surface on the LDso of DNOC and its hyperthermic effect were evaluated in male rats (Stoner 1969). The intraperitoneal LDs° of DNOC was significantly (p<0.001) reduced from 24.8 to 26.2 mg/kg to 14 mg/kg DNOC when DNOC was given 1.5- 24 hours after either type of nonfatal injury. The authors concluded that the toxicity of DNOC was increased by previous trauma. These investigators proposed that this interaction was associated with sequential blocking of the tricarboxylic acid cycle with inhibition of citrate synthetase reaction during the early part of the response to the injury. Because DNOC acts as an uncoupler of oxidative phosphorylation, less ATP is produced. Therefore, the effects of trauma will be enhanced by an uncoupling agent such as DNOC. 

NS), respectively, cytochrome oxidase decreased 9%(NS). Treatment-related effects in wmn muscle citrate synthetase decreased 19% (NS), cytochrome oxidase increased 23% (NS), muscle lactate and lactic acid dehydrogenase affected. Subjects reported no adverse health effects after exposure. 

Citrate Synthetase Isocitrate Lyase Malate Synthetase Malate Dehydro- genase Succinate Dehydro- genase Fumarase... 

Following are a set of assay conditions for marker enzymes of mitochondria (citrate synthetase, malic dehydrogenase, fumarase, and succinate dehydrogenase) and glyoxysomes (citrate synthetase, malate synthetase, and malic dehydrogenase). Some or all of these activities may be assayed across the density gradient. Their quantitative distribution is shown in Table 9-2. 

Citrate Synthetase. The assay method is modified from that of Srere, Brazil, and Gonen. The reaction mixture contains, in a volume of 1.3 ml, 7.7 X 10 M Tris (adjusted to pH 8.0), 1.5mM 5,5 -dithiobis-(2-nitrobenzoic acid), l.lmM MgCU, 2.6mM oxalacetate, 1.8 x lO M acetyl-CoA, and from 2 to 25 ig of protein, depending on which fraction of gradient is being assayed. The assay is initiated by the addition of the... 

The cleavage of citrate to acetate and oxalacetate has a AG of — 680 cal/mole. The of the citrate synthetase reaction is 3.2 X 10. From this information, calculate the standard free energy of hydrolysis of acetyl-S-CoA and the Ai, for the hydrolysis. 

The rate at which the citric acid cycle operates is precisely adjusted to meet cellular needs for ATP. One important control point is the very first step in the cycle, the synthesis of citrate. The catalyst for this reaction, citrate synthetase, is an allosteric enzyme that is... 

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