• BMB Reports
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• v.28 no.6
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• pp.471-476
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• 1995

Acetolactate synthase (ALS) catalyzes the first common step in the biosynthesis of valine, leucine, and isoleucine. ALS is the target enzyme for several classes of structually diverse herbicides. We have synthesized 4,6-dimethoxypyrimidine derivatives as ALS inhibitors, and their inhibitory activities on barley ALS were determined. $IC_{50}$ values for the derivatives are 0.2~200 ${\mu}m$. K11570, the most potent ALS inhibitor with $IC_{50}$ of 0.2 ${\mu}m$, showed mixed-type inhibition with respect to substrate pyruvate, and the progress curves for ALS inhibition by K11570 indicated that the amount of inhibition increased with time. Inhibition-competition experiments were carried out and indicated that three different classes of inhibitors, K11570, a sulfonylurea Ally, and leucine, bind to ALS in a mutually exclusive manner. Chemical modification of tryptophanyl and tyrosyl residues of ALS decreased the sensitivity of ALS to K11570, while cysteine modification did not affect the sensitivity. These results suggest that tryptophanyl and tyrosynyl residues are probably located at or near the inhibitor binding site.

• The Korean Journal of Pesticide Science
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• v.5 no.3
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• pp.41-46
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• 2001

This study was conducted to develop a high throughput system for screening acetolactate synthase(ALS) inhibitors, and to detect basic mother molecules for developing new novel herbicide candidates. The high throughput screening (HTS) method using 96-well plate and microplate reader was developed. This method is 8 times more effective than basic technique in one cycle per person. Futhermore, considering for less than 1/10 volume of materials required for ALS test and enzyme kinetics with 16 times faster speed compared to those of former procedure, this HTS method has more than 100 times higher efficacy than basic system in a consecutive procedure. We discovered 11 new ALS inhibitors such as 2-oxoglutaric acid, aminooxyacetic acid, azelaic acid, citric acid, cyanuric fluoride, itaconic acid, malonic acid, niclosamide, oxalic acid, glyoxylic acid, and suramin from 107 commercial plant-specific inhibitors using this technique. We hope these results might be useful to discover lead compounds for developing new novel herbicide candidate.

• Korean Journal of Weed Science
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• v.16 no.2
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• pp.122-131
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• 1996

The inhibition characteristics of chlorsulfuron [CHL, 2-chloro-N-[{ (4-methoxy-6-methyl-1,3,5-triazin-2-yl)amino}carbonyl]benzenesulfonamide] and imazaquin [IMA, 2-{4,5-dihydro-4-methyl-4-(1-methy-lethyl)-5-oxo-1H-imidazol-2-yl}-3-quinolinecarboxylic acid] on acetolactate synthase(ALS) activity of corn plants were investigated. CHL and IMA rapidly inhibited ALS activity of corn plants in vitro. Their $I_{50}$ values for ALS activity were 100nM and $5{\mu}M$, respectively, indicating that CHL had 50 times more inhibitory effect on ALS activity than IMA. The first applied herbicide had a dominant inhibitory effect on ALS activity when the two herbicides were applied sequentially. Branched-chain amino acids, valine(Val), leucine(Leu), and isoleucine(Ile) showed a feedback inhibition on ALS activity ; Val or Leu had a more inhibitory effect on ALS activity than Ile. Branchedchain amino acids and CHL or IMA exhibited an additive effect on inhibiting ALS activity. This suggests that branched-chain amino acids inhibit ALS activity by a different mechanisms) from that of CHL or IMA. Apparent ALS activity, which was measured on the basis of the conversion of pyruvate to acetolactate, was decreased by the addition of 2-ketobutyrate into the ALS reaction mixture in a concentration-dependent manner. In addition, kinetic studies revealed that CHL acts as a noncompetitive inhibitor, while IMA acts as an uncompetitive inhibitor to ALS with respect to pyruvate.

• Journal of the Korean Chemical Society
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• v.41 no.6
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• pp.304-312
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• 1997

Acetolactate synthase(ALS) is the common enzyme in the biosynthetic of valine, leucine, and isoleucine, and is the target of several classes of structually unrelated herbicides, including sulfonylureas, imidazolinones, and triazolopyrimidines. In an effort to develop new and desirable herbicides, we have synthesized 4,6-dimethoxypyrimidine derivatives, and examined their inhibitory activities on pea ALS. The most active compound was found to be K11570 and $IC_{50}$ value for K11570 was 0.2 ${\mu}M.$ The inhibition of pea ALS by K11570 was biphasic, showing increased inhibition with incubation time. The K11570 showed mixed-type inhibition with respect to substrate pyruvate. Dual inhibition analysis of K11570 versus sufonylurea herbicide Ally and feedback inhibitor leucine revealed that three inhibitors were competitive for binding to ALS. The arginine modified enzyme showed decreased inhibition by K11570, sufonylurea Ally, and leucine, in constrast to, tryptophan modification did not affect on the sensitivity of ALS to the inhibitors.

• Korean Journal of Weed Science
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• v.17 no.1
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• pp.66-70
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• 1997

LGC-40863(proposed common name ; pyribenzoxim), (benzophenone O-[2,6-bis[(4,6-dimethoxy-2-pyrimidinyl)oxy]benzoyl]oxime) is a new rice herbicide being developed by LG Chemical Ltd. The herbicide is highly selective between rice(Oryza sativa L.) and weeds including barnyardgrass (Echinochloa crus-galli(L.) P. Beauv.), and assumed to inhibit acetolactate synthase(ALS ; EC 4.1.3.18) because other structurally related herbicides inhibit the enzyme. To know inhibitory activity and the mode of inhibition of LGC-40863, $I_{50}$(concentration inhibiting ALS activity by 50%) and inhibition kinetics were investigated using ALS extracted from rice and barnyardgrass. $I_{50}$ values of LGC-40863 were 14 and 16mM in rice and barnyardgrass, respectively. In contrast to imazapyr(2-[4,5-dihydro-4-mythyl-4-(1-methylethyl)-5-oxo-1H-imidazol-2-yl]-3-pyridine-carboxylic acid) which showed an uncompetitive inhibition pattern, LGC-40863 was a noncompetitive inhibitor to ALS with respect to pyruvate similar to chlorsulfuron(2-chloro-N-((4-methoxy-6-methyl-l,3,5-triazin-2-yl) aminocarbonyl)benz-enesulfonamide) in both plants.

• Journal of the Korean Chemical Society
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• v.42 no.3
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• pp.306-314
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• 1998

Acetolactate synthase (ALS) is the common enzyme in the biosynthesis of branched chain amino acids, Val, Leu, and Ile in bacteria, yeast, and higher plants. The enzyme is target site of several classes of structually diverse herbicides, including the sulfonylureas, the imidazolinones, the triazolopyrimidines, and the primidyl-oxy-benzoates. We have synthesized new triazolopyrimidine (TP) derivatives, and determined their inhibitory activities on barley ALS. $lC_{50}$ values for the active compounds were 3.2 nM-0.62 mM, and some of them appeared to be potent inhibitors. The progress curves for inhibition of ALS by TP4, a representative derivative, indicated that the extent of inhibition increased with incubation time. The inhibition of ALS by TP4 showed mixed-type inhibition with respect to pyruvate. Dual inhibition analyses of TP4 versus imidazolinone Cadre and feedback inhibitor Leu suggested that three different classes of inhibitors bind to ALS in a mutually exclusive manner. Chemical modification of tyrosyl residues of ALS decreased sensitivity of ALS to TP4, while modification of tryptophan and cysteine did not affect the sensitivity.

• Weed & Turfgrass Science
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• v.3 no.2
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• pp.110-113
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• 2014

The continuous use of acetolactate synthase (ALS) and acetyl-CoA carboxylase (ACCase) inhibitors has led to the selection of herbicide resistant barnyardgrass populations in direct-seeded rice fields of Korea. This study was conducted to identify herbicide resistant barnyardgrass biotypes and to determine the cross- and multiple-resistance of them. 25% of the population collected from Taeahn was partially resistant to ACCase inhibitors and 22% collected from Kimjae were partially resistant to ALS inhibitors. However, 8.2% of the population from both sites was resistant to ALS and ACCase inhibitors. Resistance to sulfonylurea herbicide, flazasulfuron was identified from two barnyardgrass accessions collected from both Taeahn and Kimjae. One barnyardgrass accession from both sites was resistant to ACCase inhibitor, sethoxydim. The cross-resistance to ALS inhibitors was identified at one barnyardgrass accession from Taeahn and at two accessions from Kimjae. Further, crossresistance to ACCase inhibitors was also identified at barnyardgrass accessions from Taeahn and Kimjae. Multiple-resistance to flazasulfuron and sethoxydim was determined at four barnyardgrass accessions from Taeahn and at six accessions from Kimjae. Therefore, the herbicide mixture and sequences within a growing season or the herbicide rotation with different modes of actions across growing seasons are recommended to control herbicide-resistant barnyardgrass in infested fields.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.47 no.4
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• pp.328-334
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• 2002

A Lindernia dubia (L.) Pennell var. dubia accession from Jeonnam province, Korea was tested for resistance to sulfonylurea (SU) herbicides, imazosulfuron and pyrazosulfuron-ethyl in whole-plant response bioassay. The accession was confirmed resistant to both herbicides. The $GR_{50}$ (herbicide concentration that reduced shoot dry weight by 50%) values of resistant accession were 264 and 19 times higher to imazosulfuron and pyrazosulfuronethyl, respectively, than that of the standard susceptible accession. The surviving resistant L. dubia after pyrazosulfuron-ethyl + molinate application can be controlled by sequential applications of soil-applied herbicides, butachlor, dithiopyr, pyrazolate, and thiobencarb and foliar herbicides, bentazon. Sulfonylurea-based mixtures such as mixtures of azimsulfuron + anilofos, bensulfuron-methyl + oxadiazon, pyrazosulfuron-ethyl + fentrazamide, and pyrazosulfuron-ethyl + anilofos + carfentrazon can also be used to control the surviving resistant L. dubia. However, use of these mixtures should be restricted to a special need basis. Thus, we suggest that sequential applications of non-SU-based mixtures such as butachlor + pyrazolate and MCPB + molinate + simetryne be used to control the surviving resistant L. dubia after SU herbicide applications. Rice yield was reduced 24 % by resistant L. dubia that survived after the pyrazosulfuron-ethyl + molinate application compared with pyrazolate + butachlor in transplanted rice culture. In vitro ALS activity of the resistant biotype was 40 and 30 times more resistant to imazosulfuron and pyrazosulfuron-ethyl, respectively, than the susceptible biotype. Result of in vitro ALS assay that the resistance mechanism of L. dubia to SU herbicides may be due, in part, to an alteration in the target enzyme, ALS.

• Journal of the Korean Chemical Society
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• v.39 no.6
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• pp.459-465
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• 1995

Acetolactate Synthase (ALS) was partially purified from the yeast and its basic biochemical studies were carried out. Yeast was grown in the minimum media containing 0.5% glucose, 51 mM $K_2HPO_4$, 22 mM $KH_2PO_4$, 8 mM $(NH_4)2SO_4,\;0.4\;m M\;MgSO_4$ for 18 hours at 37 $^{\circ}C$. The cell was ruptured in the buffer (20 mM phosphate buffer pH 7.0, 0.1 mM TPP, 0.5 mM DTT, 1 ${\mu}M$ FAD, and 1 mM MgCl_2$) following an overnight suspension. The supernatant fraction was collected from $10,000{\times}g$ and the enzyme was further purified by ammonium sulfate fractionation, DEAE-Sephacel chromatography and leucine-agarose chromatography. The enzyme activity was measured under the various conditions by the function of protein concentration, time, temperature, pH, and substrate. The optimum temperature was found to be 50$^{\circ}C$, optimum pH 8.0∼8.5. The kinetic parameters, $K_m\;and\;V_{max}$ were 8.4 mM and 17.9 nmol/mg/min respectively. Stability of the enzyme was studied with ethylene glycol and glycerol added to the enzyme solution. Both ethylene glycol and glycerol improved the enzyme stability up to 50%. The study of feedback inhibition showed that valine was a strong inhibitor while leucine was a weak inhibitor.

• Weed & Turfgrass Science
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• v.6 no.4
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• pp.345-349
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• 2017

Herbicides are important weed control tools for increasing crop yields and the efficiency of crop production. As the use of herbicides increases, the occurrrence of herbicide-resistant weeds has been an increaing problem. In Korea, since the first occurrence of acetolactate synthase (ALS) inhibitor resistant Monochoria korsakowii was reported in the Seosan reclaimed paddy field in 1998, resistance has been reported in 14 weed species, including Echinochola spp. and their populations are gradually increasing. The objective of this study is to investigate the nationwide occurrence of ALS and Acetyl-CoA Carboylase inhibitor resistant Echinochloa spp. in Korea. In 2013, 2014, and 2015, we collected 594 accessions of Echinochloa spp. in Korean rice fields except for Jeonnam and Chungbuk provinces. They were then treated with the recommended rates of penoxsulam and cyhalofop-butyl. We harvested seeds from 45 accessions of E. oryzicola in the case of cyhalofop-butyl treatment. Also, 44 and 46 accessions of E. oryzicola and E. crus-galli survived and their seeds were harvested after penoxsulam treatment. Twenty accessions of E. oryzicola survived from both herbicides inferring possible multiple resistance. Two accessions out of 20 inferred from possible multiple resistance survived after cyhalofop-butyl treatment at a dose of $500ga.i.ha^{-1}$. Seeds of herbicide resistant populations will be provided and utilized for further research.