Inhibitors of Branched Chain Amino Acid Biosynthesis
Branched Chain Amino Acid Biosynthesis
Branched chain amino acid production is important for several reasons. Valine, leucine and isoleucine represent 3 of the 10 essential amino acids needed for human nutrition. These amino acids are critical for protein synthesis and normal plant growth, while also providing precursors for a number of secondary metabolites such as; cyanogenic glycosides, glucosinolates, and acyl-sugars. Branched chain amino acid production also represents a commercially important target for a variety of low-use-rate herbicides.
Branched chain amino acid production starts with two different substrates. Valine and leucine production starts with pyruvate derived directly from photosynthesis, while isoleucine production starts with the deamination of threonine by threonine deaminase to form 2-ketobutyrate. From this point, valine and isoleucine production share four enzymes, each with two substrates. While there are three enzymes shared in the production of all three amino acids, the first shared enzyme, acetolactate synthase [ALS; EC220.127.116.11(1)] is the enzyme that will be the focus of this lesson. ALS is also referred to acetohydroxyacid synthase or AHAS.
ALS is a tetramer made up of four identical subunits and it belongs to a larger group of enzymes, called a superfamily, that utilize thiamine pyrophosphate (TPP) as part of the catalytic process. ALS requires two additional cofactors, Mg2+ and flavin dinucleotide (FAD). The Mg2+ appears to play an important role in binding TPP along with two highly conserved amino acids of ALS. Because FAD does not appear to participate directly in the activity of this enzyme it has been suggested that it might be a hold over from some ancestral form of the enzyme. Each of the four identical subunits consists of three identifiable regions called domains. In general terms, these domains are a combination of ß sheets and alpha helices (See Figure 1).
The reaction carried out by ALS is the synthesis of either 2-acetolactate or 2-acetohydroxybutyrate. Acetolactate is produced by the combination of two pyruvate molecules, while acetohydroxybutyrate is formed by the combination of 2-ketobutyrate with pyruvate (see animation). Plant ALS is feedback inhibited by valine and leucine and these amino acids appear to act synergistically. This could be the result of allosteric regulation, meaning that the binding of one feedback inhibitor facilitates the binding of the second.
The gene that encodes for ALS is found in the plant cell nucleus; however, the pathway is located in the chloroplast. That means that somehow the ALS enzyme must be transported from the nucleus to the chloroplast. Since the major precursor needed to start the branched chain amino acids biosynthesis is a product of photosynthesis it makes sense energetically to transport the enzyme to the chloroplast. This is accomplished in the same manner as with EPSP synthase. The gene encoding ALS has a “leader sequence” that directs the enzyme to the chloroplast where it is cleaved to produce an active ALS enzyme. The fact that the ALS gene is nuclear encoded has significant implications with regard to the spread of the resistance gene. This will be discussed in other parts of this lesson.
Development of this lesson was supported in part by Cooperative State Research, Education, & Extension Service, U.S. Dept of Agriculture under Agreement Number 98-EATP-1-0403 administered by Cornell University and the American Distance Education Consortium (ADEC). Any opinions, findings, conclusions or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture.
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