Oxaloacetic acid synthesis, catalysed

Figure 11.3 Mechanism of transfer of acetyl-CoA out of the mitochondrion. In the mitochondrion, acetyl-CoA reacts with oxaloacetate to form citrate, which is transported across the mitochondrial inner membrane. In the cytosol, citrate is split to re-form citrate and oxaloacetate, catalysed by citrate lyase. It has been shown that inhibition of citrate lyase inhibits fatty acid synthesis.

The biological effect of (-)-HCA stems from the inhibition of extramitochondrial cleavage of citrate to oxaloacetate and acetyl-CoA catalysed by ATPicitrate lyase. This limits the availability of acetyl-CoA units required for fatty acid synthesis and lipogenesis (Sullivan, 1984). The inhibition of ATP cit-rate lyase by (-)-HCA leads to less dietary carbohydrate utilization for the synthesis of fatty acids, resulting in more glycogen storage in the liver and muscles. Many in vitro... 

In animals and fungi there is a similar dichotomy. NADPH can be generated by cytosolic malic enzyme which catalyses the reaction malate + NADP+ — pyruvate + COg + NADPH. Cytosolic malate derives from the following successive reactions the pyruvate/ citrate shuttle on the mitochondrial inner membrane takes pyruvate to the mitochondrion in exchange for citrate cytosolic ATP citrate lyase catalyses ATP + citrate + CoA-SH —> acetylCoA (CH3CO-S-C0A) + oxaloacetate and cytosolic malate dehydrogenase, which catalyses NADH + oxaloacetate NAD+ + malate. This scheme provides both acetylCoA and NADPH for subsequent long chain fatty acid synthesis (see section on Fatty acid synthesis ). 

The ketoacyl-ACP is then reduced to yield a hydroxyl group. In turn, this is dehydrated to yield a carbon-carbon double bond, which is reduced to yield a saturated fatty acid chain. Thus, the sequence of chemical reactions is the reverse of that in P-oxidation (section 5.5.2). For both reduction reactions in fatty acid synthesis, NADPH is the hydrogen donor. One source of this NADPH is the pentose phosphate pathway (section 5.4.2) and the other is the oxidation of malate (arising from oxaloacetate) to pyruvate, catalysed by the malic enzyme (see Figure 5.27). 

Pyruvate carboxylase catalyses the conversion of pyruvate to oxaloacetate using ATP and CO2. This is an important reaction both for gluconeogenesis (to bypass the pyruvate kinase reaction) and also for the normal function of the citric acid cyde. For the citric acid cycle to begin, one molecule each of oxaloacetate and acetyl CoA is required. If there is a shortage of oxaloacetate, the balance is restored by the action of pyruvate carboxylase. If, for example, oxaloacetate has been removed from the cycle to enter the amino acid synthesis pathways, it can simply be regenerated fiom pyruvate (31b). 

We mentioned the citric acid cycle earlier but we have not so far discussed the chemistry involved. The citric acid cycle allows metabolism to shunt carbon atoms between small molecules, and the key step is the synthesis of citric add from oxaloacetate and acetyl CoA. The reaction is essentially an aldol reaction between the enol of an acetate ester and an electrophilic ketone, and the enzyme which catalyses the reaction is known as citrate synthase. 

On the other hand, a-transaminases have been used extensively in the production of amino acids through kinetic resolution and asymmetric synthesis. While many studies rely on the use of an excess of cosubstrate to drive the reaction to completion, some multienzymatic approaches have been developed as well. As an example, aspartate has been used as an amino donor in a multienzymatic synthesis of L-2-aminobutyrate from L-threonine (Scheme 4.8). ° The rather complex multistep sequence started with the in situ formation of 2-ketobutyrate from L-threonine catalysed by threonine deaminase (ThrDA) from E. coli. A tyrosine transaminase (lyrAT) from E. coli converted 2-ketobutyrate and L-aspartie acid to L-2-aminobutyrate and oxaloacetate, which spontaneously decarboiq lated to give pyruvate. Since the... 

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