Imidazolinones, enzyme inhibition

The aromatic ring component illustrates the relative contributions of enzyme inhibition and physicochemical properties to herbicidal activity. The benzene imidazolinones are approximately ten-fold more potent than the corresponding pyridine derivatives as enzyme inhibitors but are less potent as herbicides. 

Herbicides control weeds and are the most widely used class of pesticides. The latest US EPA data show that some 578 million pounds of herbicides were used in the United States in 1997 and accounts for some 47% of pesticides used. This class of pesticide can be applied to crops using many strategies to eliminate or reduce weed populations. These include preplant incorporation, pre- and postemergent applications. New families of herbicides continue to be developed, and are applied at low doses, are relatively nonphytotoxic to beneficial plants and are environmentally friendly. Some of the newer families such as the imidazolinones inhibit the action of acetohydroxyacid synthase that produces branched-chain amino acids in plants. Because this enzyme is produced only in plants, these herbicides have low toxicities to mammals, fish, insects, and birds. 

ALS was isolated from barley seedlings as a 0-33% Ammonium Sulfate precipitate and examined for inhibition by TP. It is apparent from Figure 5 that the enzyme is very sensitive to the compound. The 1(50) value (concentration required for 50% inhibition) was calculated to be 0.047 uM. This value is within the range reported for CS tested against ALS from different species (19). Imidazolinones are less potent with 1(50) values in the range 2-12 uM (26). ALS isolated from several species and their 1(50) values for TP is shown in Table I. 

Lolium biotypes exist which have resistance to the sulfonylurea herbicides chlorsulfuron and metsulfuron methyl (4). The biotype used in the studies presented here is resistant to both these sulfonylurea herbicides. Sulfonylurea herbicides inhibit the chloroplastic enzyme acetolactate synthase (ALS), also known as acetohydroxyacid synthase (AHAS) (16). Inhibition of this enzyme results in disruption of the synthesis of the branched-chain amino acids valine and isoleucine (161. The imidazolinone herbicides also inhibit ALS Q2). In some species auxins can protect against chlorsulfuron inhibition (S. Frear, USDA North Dakota, personal communication) the mechanistic basis for this protection is not known. We have measured the ALS activity in the resistant and susceptible Lolium and have also checked for any induction of ALS activity following treatment with the sulfonylurea herbicide chlorsulfuron. 

Identification of the mode of action of the imidazolinones occurred while resistant cell lines were being isolated. Imidazolinones inhibit acetohydroxyacid synthase (AHAS EC 4.1.3.18), the first enzyme in the pathway of branched chain amino acid synthesis (8). Imidazolinone-resistant cell lines provide proof that inhibition of AHAS is the site of action of the imidazolinones AHAS activities in extracts from resistant corn cell lines are highly resistant to inhibition by imidazolinone herbicides (7). 

For example, glyphosate inhibits the enzyme, EPSP (5-enolpyruvylshikimate 3-phosphate) synthase, that catalyzes a step in the synthesis of the aromatic amino acids. Similarly, both the imidazolinones and sulfonylureas inhibit acetolactate synthase (ALS), the enzyme that catalyzes the first step in the formation of branched-chain amino acids (11). Triazine herbicides act by binding to a specific protein in the thylakoid membranes of the chloroplasts, preventing the flow of electrons and inhibiting photosynthesis (12). 

Inhibitors of Acetolactate Synthase (ALS/AHAS) The enzyme acetolactate synthase (ALS) plays in plants an essential role in branched-chain amino acid biosynthesis. In the pathway leading to valine and leucine, ALS catalyzes the formation of 2-acetolactate from two pyruvate molecules, and in the pathway to isoleucine the formation of 2-acetohydroxybutyrate from 2-ketobutyrate and pyruvate. Due to this double function the enzyme is also called with a more general term aceto-hydroxyacid synthase. ALS is inhibited by several groups of herbicides, mainly the sulfonylureas (SUs), imidazolinones (IMIs), triazolopyrimidines (TPs), pyrimidinylthiobenzoates(PTBs) and sulfonylaminocarbonyltriazolinone (SCTs) (see Chapter 2.1, M. E. Thompson). 

These imidazolinone tolerance traits in different crops were developed by various methods. These methods included tissue culture selection (maize), pollen mutagenesis (maize), microspore selection (canola), seed mutagenesis (wheat and rice) and incorporation of resistance trait from a weedy relative (sunflower). Details of these methods have been previously reviewed [31, 32]. In all of these cases, the basis of tolerance is due to the presence of an altered form of AHAS that is resistant to inhibition by imidazolinones. The resistant enzyme is produced due to a single base pair change in the gene encoding the large subunit... 

Imazaquin, an example of the imidazolinone class of herbicides, is particularly selective to soybeans. These compounds have a similar mode of action to that of the sulfonylurea herbicides, which will be discussed later. They block branched chain amino acid biosynthesis by inhibition of the enzyme acetolactate synthase (ALS). 

ALS shows a high degree of primary sequence homology with pyruvate carboxylase and pyruvate oxidase the ubiquinone cofactors of pyruvate oxidase inhibit ALS, and it has been proposed that the ubiquinone-binding site of the ancestral enzyme also is the site of both SMM and imidazolinone binding. Recent evidence suggests, however, that these two types of herbicide interact differently with ALS (a) imidazolinones cause a rapid decrease in the levels of extractable ALS activity in maize, whereas SMM does not and can protect the ALS activity from this in vivo effect of imidazolinones and (b) not all imidazolinone-tolerant cell lines are insensitive to sulfonylureas. Sulfonylureas and imidazolinones do not, however, show synergistic inhibition of maize ALS in vitro. ... 

Acetolactate synthase (ALS) is the enzyme target site of the sulfonyl-ureas. In common with the imidazolinone aryl carboxylates, these herbicides inhibit valine and isoleucine biosynthesis. The imidazolinones are exemplified by Assert (8), which consists of a mixture of m- and p-isomers. The selectivity of ALS inhibitors invariably can be accounted for by differential metabolism or uptake or related phenomena rather than by any significant inherent difference in the properties of the ALS enzymes of crop and weed species. Assert is another example of a herbicide activated in plants by deesterification to the phytotoxic acid, and susceptible species such as Avena fatua (wild oat) are unable to metabolize the molecule further. Facile ring-methyl hydroxylation to the m- and p-benzyl alcohols, however, followed by glycosylation confers tolerance to maize and wheat." ... 

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