An Intermediate Trapped by Maleimides in a Pyridoxal-Phosphate Potentiated Enzymatic Elimination Reaction

Flavin, Martin; Slaughter, Clarence

doi:10.1021/bi00895a004

Cited by 39 publications

(23 citation statements)

References 12 publications

(21 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Flavjn and coworkers have shown that when N-ethylmaleimide is added to reaction mixtures containing pyridoxal-dependent enzymes catalyzing 8-or y-elimination reactions and the appropriate substrates, a new keto acid is formed which contains the N-ethylmaleimide moiety [30,31,34]. N-Ethylmaleimide apparently reacts with an enzyme-substrate intermediate having carbanion character a t carbon atom 3 of the substrate to form an addition product; the latter eventually splits off from the enzyme.…”

Section: Discussionmentioning

confidence: 99%

“…The Effect of N-Ethylmaleimide on the @-Elimination Reaction Flavin and coworkers have recently proposed the use of N-ethylmaleimide as a trapping agent for carbanion intermediates in some enzymatic 8or y-elimination reactions [30,31]. I n the present study an experiment was carried out in which N-ethylmaleimide was added to incubation mixtures of /3-chloroglutamic acid and glutamate-aspartate transaminase.…”

Section: Transamination Between P-chloroglutamate and Keto Acidsmentioning

confidence: 99%

See 1 more Smart Citation

The Reaction of β‐Chloroglutamic Acid with Glutamate‐Aspartate Transaminase

Manning

Khomutov

Fasella

1968

European Journal of Biochemistry

View full text Add to dashboard Cite

Porcine glutamate-aspartate transaminase catalyzes a @-elimination reaction with both the threo-and erythro-isomers of 8-chloroglutamate ; chloride, ammonia, and a-ketoglutarate are formed in equimolar amounts. The latter product was characterized as the 2,4-dinitrophenylhydrazone and by catalytic hydrogenation of this derivative to glutamic acid.The 8-elimination reaction is catalyzed by highly purified glutamate-aspartate transaminase ; the reaction is not catalyzed by the phosphopyridoxamine form of the enzyme, the apoenzyme, or by free pyridoxal 5'-phosphate. There is no detectable transamination between B-chloroglutamate and either oxaloacetate or a-ketoglutarate. Incubation of either the holoenzyme or the apoenzyme with p-chloroglutamate does not result in any detectable loss of enzyme activity. I n the presence of N-ethylmaleimide much less a-ketoglutarate than ammonia is formed; this is consistent with the idea that a reactive carbanion intermediate reacts with N-ethylmaleimide.The novel 8-elimination reaction with glutamate-aspartate transaminase demonstrated here supports the idea that in some instances the specificity of an enzymatic reaction can be determined by the structure of the substrate.Glutamate-aspartate transaminase catalyzes the reversible transfer of an amino group fromL-aspartate to a-ketoglutarate forming oxaloacetate and L-glutamate; other amino acids can also undergo transamination with this enzyme although they are less efficient substrates than the glutamate-aspartate pair [I]. There has been considerable interest in the reaction of substrate analogues with glutamate-aspartate transaminase in an effort to elucidate some of the intermediate steps of the reaction. Thus, in a study of the reaction between the transaminase and a-methylaspartate (which lacks an a-hydrogen atom and hence cannot undergo transamination) two new absorption bands were observed, one at 430 mp and the other at 360 mp [2]. It was proposed that these are due to coenzyme-substrate aldimine derivatives analogous to those formed with the glutamateaspartate substrate pair before the loss of the a-hydrogen atom. When erythro-/3-hydroxyaspartic acid is incubated with glutamate-aspartate transaminase there is a dramatic increase in the absorption a t 490 mp; this is followed by a slow increase in the absorption a t 330 mp (attributed to the phosphopyridoxamine enzyme) and a corresponding formation Enzymes. Glutamate-aspartate transaminase, or L-aspartate: 2-oxoglutarate aminotransferase (EC 2.6.1.1) ; glutamic dehydrogenase, or L-glutamate : NAD oxidoreductase (deaminating) (EC 1.4.1.2) ; malic dehydrogenase, or L-malate: NAD oxidoreductase (EC 1.1.1.37); glutamic decarboxylase, or L-glutamate: 1-carboxy-lyse (EC 4.1.1.15).18 European J. Biochem., Vol. 5 of keto acid. However, little, if any, of the intcrmediate absorbing a t 490mp is detected when threo-@-hydroxyaspartate is the substrate. It has been suggested that the species absorbing a t 490 mp is a semiquinoid coenzyme-substrate derivative [3]. The reason for the d...

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Transamination Between P-chloroglutamate and Keto Acidsmentioning

confidence: 99%

The Reaction of β‐Chloroglutamic Acid with Glutamate‐Aspartate Transaminase

Manning

Khomutov

Fasella

1968

European Journal of Biochemistry

View full text Add to dashboard Cite

show abstract

“…These findings suggest that IlvA orthologs from S. enterica and E. coli either do not catalyze the protonation of 2-aminocrotonate or, if 2-aminocrotonate protonation is enzyme catalyzed, the imine product can be released from the active cite and undergo tautomerization back to 2-aminocrotonate. IlvA can also use L-serine as a substrate, which generates 2AA, a reactive enamine with a half-life of less than 3 min in aqueous solution ( 9 , 10 ). Significantly, 2AA damages some PLP-dependent enzymes by a covalent modification which involves forming a pyruvate–PLP adduct and thus inactivating the enzyme ( 11 , 12 ) ( Fig.…”

mentioning

confidence: 99%

2-Aminoacrylate stress damages diverse PLP-dependent enzymes in vivo

Shen

Borchert

Downs

2022

Journal of Biological Chemistry

View full text Add to dashboard Cite

“…This suggested that L. cymbulifera might synthesize phytotoxic terpenoids continuously, by which they accumulate in surrounding rhizosphere soil and reach an effective concentration. Studies have shown that compound 3 might interfere with the enzymes involved in amino acid metabolism by reaction with pyridoxal at room temperature without any catalyst, due to the presence of its electron-rich tri-substituted furan ring (Iida et al, 2007; Torihata and Kuroda, 2008). Similarly, in our study, the phytotoxic activities of compound 3 mainly depend on the co-existence of the tri-substituted furan ring and the OH-10.…”

Section: Resultsmentioning

confidence: 99%

Phytotoxic Terpenoids from Ligularia cymbulifera Roots

et al. 2017

View full text Add to dashboard Cite

Ligularia cymbulifera is one of the predominant species in the Hengduan Mountains, China, and has led to a decrease in the amount of forage grass in this area. However, little is known about the mechanism behind its predominance. In this study, two novel eremophilane sesquiterpenes, ligulacymirin A and B (1 and 2), together with seven other known terpenoids (3–9), were isolated from the roots of L. cymbulifera. The structures of 1 and 2 were determined by spectroscopic methods and single-crystal X-ray diffraction. Each compound showed phytotoxic activities against Arabidopsis thaliana, and each was detected and identified in rhizosphere soil by UHPLC-MS. Compound 3 was the most potent phytotoxin, showing remarkable inhibition against both seedling growth (EC50 = 30.33 ± 0.94 μg/mL) and seed germination (EC50 = 155.13 ± 0.52 μg/mL), with an average content in rhizosphere soil of 3.44 μg/g. These results indicate that terpenoids in L. cymbulifera roots might be released as phytotoxins in rhizosphere soil to interfere with neighboring plants.

show abstract

An Intermediate Trapped by Maleimides in a Pyridoxal-Phosphate Potentiated Enzymatic Elimination Reaction

Cited by 39 publications

References 12 publications

The Reaction of β‐Chloroglutamic Acid with Glutamate‐Aspartate Transaminase

The Reaction of β‐Chloroglutamic Acid with Glutamate‐Aspartate Transaminase

2-Aminoacrylate stress damages diverse PLP-dependent enzymes in vivo

Phytotoxic Terpenoids from Ligularia cymbulifera Roots

Contact Info

Product

Resources

About