Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

---

Query: KEGG:D02011 (FAD)
5,530 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The gene encoding the valine-resistant and FAD-independent acetolactate synthase of Klebsiella pneumoniae was isolated and expressed in Escherichia coli. The nucleotide sequence of this gene was determined and it exhibited an open reading frame of 1680 bp in length. In vivo expression of the acetolactate synthase-encoding gene in E. coli revealed a single 60-kDa protein which is consistent with the molecular weight calculated from the deduced amino acid sequence of the gene product. The gene product shares about 20-30% homology with the acetolactate synthases of E. coli, yeast and higher plants.
...
PMID:Cloning, sequencing and heterologous expression of a Klebsiella pneumoniae gene encoding an FAD-independent acetolactate synthase. 164 3

The yeast ilv2 gene, encoding acetolactate synthase, was subcloned in an Escherichia coli expression vector. Although a major part of the acetolactate synthase synthesized by E. coli cells harbouring this vector was packaged into protein inclusion bodies, we used these recombinant E. coli cells to produce large quantities of the yeast enzyme. The yeast acetolactate synthase was purified to homogeneity using first streptomycin and ammonium sulfate precipitations, followed by T-gel thiophilic interaction, Sephacryl S-300 gel filtration, Mono Q anion exchange, and Superose 12 gel filtration chromatography. SDS/PAGE and gel filtration of the purified enzyme showed that it is a dimer composed of two subunits, each with the molecular mass of 75 kDa. The purified yeast acetolactate synthase was further characterized with respect to pH optimum, dependence of the substrate, pyruvate, and requirements of the cofactors, thiamin diphosphate, Mg2+, and FAD.
...
PMID:Purification and properties of Saccharomyces cerevisiae acetolactate synthase from recombinant Escherichia coli. 268 71

The herbicide sulfometuron methyl (SM) inhibited growth of some methanococci. Of 28 strains tested, the growth of 7 was completely inhibited by 0.55 mM SM. Growth of an additional 14 strains was partially inhibited, and the growth of 7 strains was unaffected by this concentration of SM. In some cases, the branched-chain amino acids protected growth. Growth inhibition was correlated with the Ki for SM of acetolactate synthase (ALS). For the enzymes from bacteria representative of the sensitive, partially resistant, and resistant methanococci (Methanococcus aeolicus, Methanococcus maripaludis, and Methanococcus voltae, respectively), the Ki for SM was 0.0012, 0.34, and greater than 1.0 mM, respectively. Inhibition was uncompetitive with respect to pyruvate. Based on these observations, ALS appeared to be the major if not the sole site of action of SM in the methanococci. The sensitivity of the ALS from these three methanococci to feedback inhibition by branched-chain amino acids was also quite different. Although all three were sensitive to feedback inhibition by valine, the Ki varied 20-fold, from 0.01 to 0.22 mM. Moreover, only the ALS from M. maripaludis was sensitive to inhibition by leucine, and the Ki was 1.8 mM. The Ki for isoleucine for the ALS from both M. maripaludis and M. voltae was about 0.1 mM. The ALS from M. aeolicus was not inhibited by isoleucine. In other respects, the ALSs from the methanococci were very similar. After dialysis, thiamine pyrophosphate but not FAD and Mg2+ was required for maximal activity, and they were all rapidly inactivated by oxygen. Although the methanococcal ALSs exhibited diverse properties, the range of catalytic and regulatory properties closely resembled those of the eubacterial enzymes.
...
PMID:Sulfometuron methyl-sensitive and -resistant acetolactate synthases of the archaebacteria Methanococcus spp. 365 79

A facile purification has been devised for recombinantly produced Salmonella typhimurium acetolactate synthase isozyme II. Purification of the enzyme was made possible by determining the complex set of factors that lead to loss of enzymic activity with this rather labile enzyme. When complexed with thiamin pyrophosphate, FAD, and magnesium, acetolactate synthase is subject to oxygen-dependent inactivation, a property not shared by the enzyme-FAD complex. When divorced from all of its tightly bound cofactors, losses of the enzymic activity are encountered at low ionic strength, especially at low protein concentrations. If purified and stored as the enzyme-FAD complex, acetolactate synthase is quite stable. The enzyme is composed of two types of subunits, a result that was not anticipated from previous studies of ilvG (the gene that codes for the large subunit of acetolactate synthase). These subunits were determined to be in equal molar ratio in the purified enzyme from the distribution of radioactivity between the two subunits after carboxymethylation with iodo[14C]acetate and their respective amino acid compositions. Besides the expected ilvG gene product (59.3 kDa), purified acetolactate synthase contained a smaller subunit (9.7 kDa; designated here as the ilvM gene product). On the basis of sequence homology of the small subunit with that coded for by the corresponding Escherichia coli gene sequence [Lawther, R. P., Calhoun, D. H., Adams, C. W., Hauser, C. A., Gray, J., & Hatfield, G. W. (1981) Proc. Natl. Acad. Sci. U.S.A. 78, 922-925], it is encoded by the region between ilvG and ilvE, beginning at base-pair (bp) 1914 (relative to the point of transcription initiation).(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Purification and properties of Salmonella typhimurium acetolactate synthase isozyme II from Escherichia coli HB101/pDU9. 390 97

Two groups of enzymes are classified as acetolactate synthase (EC 4. 1.3.18). This review deals chiefly with the FAD-dependent, biosynthetic enzymes which readily catalyze the formation of acetohydroxybutyrate from pyruvate and 2-oxobutyrate, as well as of acetolactate from two molecules of pyruvate (the ALS/AHAS group). These enzymes are generally susceptible to inhibition by one or more of the branched-chain amino acids which are ultimate products of the acetohydroxyacids, as well as by several classes of herbicides (sulfonylureas, imidazolinones and others). Some ALS/AHASs also catalyze the (non-physiological) oxidative decarboxylation of pyruvate, leading to peracetic acid; the possible relationship of this process to oxygen toxicity is considered. The bacterial ALS/AHAS which have been well characterized consist of catalytic subunits (around 60 kDa) and smaller regulatory subunits in an alpha2beta2 structure. In the case of Escherichia coli isozyme III, assembly and dissociation of the holoenzyme has been studied. The quaternary structure of the eukaryotic enzymes is less clear and in plants and yeast only catalytic polypeptides (homologous to those of bacteria) have been clearly identified. The presence of regulatory polypeptides in these organisms cannot be ruled out, however, and genes which encode putative ALS/AHAS regulatory subunits have been identified in some cases. A consensus sequence can be constructed from the 21 sequences which have been shown experimentally to represent ALS/AHAS catalytic polypeptides. Many other sequences fit this consensus, but some genes identified as putative 'acetolactate synthase genes' are almost certainly not ALS/AHAS. The solution of the crystal structures of several thiamin diphosphate (ThDP)-dependent enzymes which are homologous to ALS/AHAS, together with the availability of many amino acid sequences for the latter enzymes, has made it possible for two laboratories to propose similar, reasonable models for a dimer of catalytic subunits of an ALS/AHAS. A number of characteristics of these enzymes can now be better understood on the basis of such models: the nature of the herbicide binding site, the structural role of FAD and the binding of ThDP-Mg2+. The models are also guides for experimental testing of ideas concerning structure-function relationships in these enzymes, e.g. the nature of the substrate recognition site. Among the important remaining questions is how the enzyme suppresses alternative reactions of the intrinsically reactive hydroxyethylThDP enamine formed by the decarboxylation of the first substrate molecule and specifically promotes its condensation with 2-oxobutyrate or pyruvate.
...
PMID:Biosynthesis of 2-aceto-2-hydroxy acids: acetolactate synthases and acetohydroxyacid synthases. 965 46

Acetolactate synthase (ALS) catalyzes the first common step in the biosynthesis of valine, leucine, and isoleucine. ALS is the target of three classes of herbicides, the sulfonylureas, the imidazolinones, and the triazolopyrimidines. Five mutants (W266F, W439F, W490F, W503F, and W573F) of the ALS gene from Nicotiana tabacum were constructed and expressed in Escherichia coli, and the enzymes were purified. The W490F mutation abolished the binding affinity for cofactor FAD and inactivated the enzyme. The replacement of Trp573 by Phe yielded a mutant ALS resistant to the three classes of herbicides. The other three mutations, W266F, W439F, and W503F, did not significantly affect the enzymatic properties and the sensitivity to the herbicides. These results indicate that the Trp490 residue is essential for the binding of FAD and that Trp573 is located at the herbicide binding site. The data also suggest that the three classes of herbicides bind ALS competitively.
...
PMID:Role of tryptophanyl residues in tobacco acetolactate synthase. 1033 29

Acetolactate synthase (ALS) is the common enzyme in the biosynthesis of valine, leucine, and isoleucine. The role of four cysteinyl residues in tobacco ALS was determined using site-directed mutagenesis and cysteine-specific cleavage. The C411A mutation abolished the enzymatic activity, as well as the binding affinity for the cofactor FAD. The activation constant of C411S for FAD is approximately 50-fold higher than that of wALS. The C607S mutation did not significantly affect the kinetic parameters. The IC(50) values of C411S and C607S for ALS-inhibiting herbicides are not much different from those of wALS. Two mutants, C163S and C309S, are labile and readily degraded to peptide fragments. The treatment of wALS with 2-nitro-5-thiocyanobenzoic acid, specific for cleavage of the N-terminal side of cysteine, yielded three peptides of 37.0, 22. 0, and 7.0 kDa. This fragmentation pattern is consistent with that deduced from the amino acid sequence of tobacco ALS, assuming the disulfide bond between Cys163 and Cys309. These results suggest that Cys411 is involved in the binding of FAD and that the intrachain disulfide bond between Cys163 and Cys309 plays a key role in maintaining the correct conformation of tobacco ALS.
...
PMID:Structural and functional role of cysteinyl residues in tobacco acetolactate synthase. 1081 42

Acetolactate synthase (ALS) is the common enzyme in the biosynthetic pathways leading to valine, leucine, and isoleucine in plants and microorganisms. ALS is the target site of several classes of structurally unrelated herbicides including sulfonylureas, imidazolinones, and triazolopyrimidines. To identify the residues conferring herbicide tolerance in tobacco ALS, site-directed mutagenesis for three residues, Ala121, Pro187 and Ser652, was performed. Mutant A121T showed strong resistance to Londax (a sulfonylurea) and Cadre (an imidazolinone), while mutant S652T was resistant only to Cadre. The S652N mutation abolished the binding affinity of FAD, and inactivated the enzyme. Double mutation of Ala121 and Ser652 with Thr yielded a mutant highly tolerant to Londax, Cadre, and TP (a triazolopyrimidine sulfonamide), but has enzymatic properties similar to those of wild-type. Substitution of Pro187 with Ser resulted in the enzyme highly susceptible to oxidation and fragmentation. These results suggest that two residues Ala121 and Ser652 are potent residues conferring herbicide resistance in tobacco ALS, and that double mutation of Ala121 and Ser652 by Thr can confer stronger tolerance to Londax, Cadre, and TP.
...
PMID:Amino acid residues conferring herbicide tolerance in tobacco acetolactate synthase. 1111 9

Acetolactate synthase (ALS) catalyzes the first common step in the biosynthesis of valine, leucine, and isoleucine in plants and microorganisms. ALS is the target of several structurally diverse classes of herbicides, including sulfonylureas, imidazolinones, and triazolopyrimidines. The roles of three well-conserved histidine residues (H351, H392, and H487) in tobacco ALS were determined using site-directed mutagenesis. Both H487F and H487L mutations abolished the enzymatic activity as well as the binding affinity for the cofactor FAD. Nevertheless, the mutation of H487F did not affect the secondary structure of the ALS. The K(m) values of H351M, H351Q, and H351F are approximately 18-, 60-, and fivefold higher than that of the wild-type ALS, respectively. Moreover, the K(c) value of H351Q for FAD is about 137-fold higher than that of wALS. Mutants H351M and H351Q showed very strong resistance to Londax (a sulfonylurea) and Cadre (an imidazolinone), whereas mutant H351F was weakly resistant to them. However, the secondary structures of mutants H351M and H351Q appeared to be different from that of wALS. The mutation of H392M did not have any significant effect on the kinetic parameters nor the resistance to ALS-inhibiting herbicides. These results suggest that the His487 residue is located at the active site of the enzyme and is likely involved in the binding of cofactor FAD in tobacco ALS. Mutational analyses of the His351 residue imply that the active site of the ALS is probably close to its binding site of the herbicides, Londax and Cadre.
...
PMID:Roles of histidine residues in tobacco acetolactate synthase. 1130 49

Acetolactate synthase (ALS) catalyzes the first common step in the biosynthesis of valine, leucine, and isoleucine. The ALS is the target of several classes of herbicides, including the sulfonylureas, the imidazolinones, and the triazolopyrimidines. The roles of three well-conserved lysine residues (K219, K255, K299) in tobacco ALS were determined using site-directed mutagenesis. The mutation of K219Q inactivated the enzyme and abolished the binding affinity for cofactor FAD. However, the secondary structure of the enzyme was not changed significantly by the mutation. Both mutants, K255F and K255Q, showed strong resistance to three classes of herbicides Londax (a sulfonylurea), Cadre (an imidazolinone), and TP (a triazolopyrimidine). In addition, there was no difference in the secondary structures of wALS and K255F. On the other hand, the mutation of K299Q did not show any significant effect on the kinetic properties or any sensitivity to the herbicides. These results suggest that Lys219 is located at the active site and is likely involved in the binding of FAD, and that Lys255 is located at a binding site common for the three herbicides in tobacco ALS.
...
PMID:Roles of lysine 219 and 255 residues in tobacco acetolactate synthase. 1205 19


1 2 Next >>

---