Branched-Chain Amino Acids: Valine and Leucine

Pyruvate (2-oxopropionate) and acetyl CoA provide the carbon backbone for the two ‘branched-chain’ amino acids, valine and leucine. Isoleucine is also a branched chain amino acid, similar in structure to leucine, but the carbon backbone is derived from aspartate. These three amino acids are often grouped together, because they share several common enzymes in their respective biosynthetic pathways (Bryan, 1980; Bryan, 1990). The same enzymes that convert 2-oxobutyrate to isoleucine also convert pyruvate to valine in a parallel but distinct pathway, with no sharing of intermediates. Synthesis of both amino acids, valine and leucine, begins with the formation of acetolactate from two molecules of pyruvate. Acetolactate synthase (ALS) also often denominated acetohydroxyacid synthase (AHAS), catalysing this condensation reaction, contains thiamine pyrophosphate as its prosthetic group. Acetolactate is subsequently reduced, rearranged and the release of water yields 2-oxoketoisovalerate. Finally, a transamination reaction by glutamate produces valine. The branch point in this pathway is 2-oxoketoisovalerate. In a methylation reaction from acetyl coA, isopropylmalate is formed. Isomerisation and decarboxylation produces 2-oxoisocaproate, which is then transaminated to leucine.

These pathways appear to be in the chloroplast since isolated chloroplasts can synthesise valine from 14CO2, and several enzymes of the pathway have been found in isolated chloroplasts. The synthesis of branched chain amino acids is also subject to feedback control by the end products. Isopropylmalate synthase is inhibited by leucine. The first enzyme, ALS, is inhibited by valine and leucine. The sulfonyl urea (e.g. chlorsulphurone) and imidazoline herbicides (e.g. imazethapyr) are very strong inhibitors of ALS, where they bind to the pyruvate-, respectively, 2-oxobutyrate-binding site and antisense mediated repression of ALS also proved ALS to be essential for plant growth and survival (Höfgen et al., 1995). However, upon overexpression of the enzyme there was no increase in the soluble level of valine, leucine and isoleucine indicating that the enzyme is not rate-limiting the biosynthetic pathway (Smith et al., 1988).

Aspartate-Derived Amino Acid Biosynthesis: Lysine, Threonine, Isoleucine, and Methionine

The aspartate pathway is a highly branched pathway, leading to the synthesis of the amino acids lysine, threonine, methionine and isoleucine (Figure 27.5). This pathway is therefore subject to a complex control by enzyme feedback inhibition loops as well as transcriptional and post-transcriptional regulation of expression of genes encoding pathway enzymes (see, Galili, 1995; Saito, 2000; Galili, 2002). Aspartate is formed either through a specific transamination catalysed by glutamate-oxalacetate transaminase or by a deamination of asparagine catalysed by asparaginase. The first two reactions of the aspartate pathway are common to all of its end-product amino acids and include the synthesis of aspartic semialdehyde from aspartate, catalysed by the enzymes aspartate kinase and aspartate semialdehyde dehydrogenase. Aspartate semialdehyde is at an important branch point, since it can either be reduced to homoserine or condensed with pyruvate to give dihydrodipicolinic acid, which subsequently undergoes a series of six enzymatic reactions to yield lysine (see section on ‘Lysine Biosynthesis and Degradation’). The other branch starts with homoserine, an amino acid not found in proteins and usually not present in appreciable concentrations in plants, with the exception of peas, where it can constitute 70% of the soluble nitrogen in 1-week old seedlings. In most plants homoserine is quickly phosphorylated to O-phosphohomoserine, which represents the next metabolic branch point, since it can be converted in a three step mechanism to methionine (see section on ‘Lysine Biosynthesis and Degradation’) or in a single step to threonine by the enzyme threonine synthase. Threonine can either be used for protein synthesis or is further metabolised to isoleucine, the synthesis of which begins with the deamination and dehydratation of threonine to 2-oxobutyrate catalysed by threonine deaminase (TD). All branched chain amino acids share a number of common enzymes converting either 2-oxobutyrate to isoleucine or pyruvate (2-oxopropionate) to valine and leucine. The steps leading to isoleucine are catalysed by acetohydroxyacid synthase (ALS or AHAS) resulting in 2-acetohydroxybutyrate, acetohydroxyacid isomerase (AHAI) yielding 2,3-dihydroxy-3-methylvalerate, which is oxidised to 2-keto-3-methylvalerate through the activity of dihydroxyacid dehydratase (DHAD). Here the pathway to leucine branches out with four further steps while isoleucine is subsequently formed by transamination through a branched chain amino acid specific aminotransferase (KAAT), specific for 2-keto-3-methylvalerate leading to isoleucine and for 2-ketoisovalerate leading to valine, respectively.

As the biosynthesis of the amino acids lysine and methionine are currently the main focus of plant biotechnology a substantial body of knowledge has accumulated recently. Therefore, we devote a separate section to each of these two pathways.

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