Amino acid synthesis threonine deaminase

It was snbseqnently discovered that the first enzyme in the pathway for isoleucine synthesis, which is threonine deaminase, was inhibited by isoleucine in an extract of E. coli. No other amino acid caused inhibition of the enzyme. Threonine deaminase is, in fact, the rate-limiting enzyme in the pathway for isoleucine synthesis, so that this was interpreted as a feedback control mechanism (Fignre 3.13(a)). Similarly it was shown that the hrst enzyme in the pathway for cytidine triphosphate synthesis, which is aspartate transcarbamoylase, was inhibited by cytidine triphosphate (Fignre 3.13(b)). Since the chemical structures of isoleucine and threonine, or cytidine triphosphate and aspartate, are completely different, the qnestion arose, how does isolencine or cytidine triphosphate inhibit its respective enzyme The answer was provided in 1963, by Monod, Changenx Jacob. 

Consider, for example, the biosynthesis of the amino acids valine, leucine, and isoleucine. A common intermediate, hydroxy ethyl thiamine pyrophosphate (hydroxy ethyl-TPP Section 17.1.1). initiates the pathways leading to all three of these amino acids. Hydroxyethyl-TPP can react with a-ketobutyrate in the initial step for the synthesis of isoleucine. Alternatively, hydroxyethyl-TPP can react with pyruvate in the committed step for the pathways leading to valine and leucine. Thus, the relative concentrations of a-ketobutyrate and pyruvate determine how much isoleucine is produced compared with valine and leucine. Threonine deaminase, the PLP enzyme that catalyzes the formation of a-ketobutyrate, is allosterically inhibited by isoleucine (Figure 24.22). This enzyme is also allosterically activated by valine. Thus, this enzyme is inhibited by the product of the pathway that it initiates and is activated by the end product of a competitive pathway. This mechanism balances the amounts of different amino acids that are synthesized. 

Figure 3. Branched chain amino acid and pantothenate synthesis. TD and ALS are the enzymes threonine deaminase and acetolactate synthase, respectively.

An excellent example of allosteric regulation—the control of an allosteric enzyme—is the five-step synthesis of the amino acid isoleucine (see I Figure 10.13). Threonine deaminase, the enzyme that catalyzes the first step in the conversion of threonine to isoleucine, is subject to inhibition by the final product, isoleucine. The structures of isoleucine and threonine are quite different, so isoleucine is not a competitive inhibitor. Also, the site to which isoleucine binds to the enzyme is different from the enzyme active site that binds to threonine. This second site, called the allosteric site, specifically recognizes isoleucine, whose presence there induces a change in the conformation of the enzyme such that threonine binds poorly to the active site. Thus, isoleucine exerts an inhibiting effect on the enzyme activity. As a result, the reaction slows as the concentration of isoleucine increases, and no excess isoleucine is produced. When the concentration of isoleucine falls to a low enough level, the enzyme becomes more active, and more isoleucine is synthesized. This type of allosteric regulation in which the enzyme that catalyzes the first step of a series of reactions is inhibited by the final product is called feedback inhibition. 

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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