Affinity Labeling of the Active Center of l-Aspartate-β-decarboxylase with β-Chloro-l-alanine

Relyea, Noël M.; Tate, Suresh S.; Meister, Alton

doi:10.1016/s0021-9258(19)42913-x

Cited by 31 publications

(11 citation statements)

References 25 publications

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“…It has generally been assumed, without any real proof, that inactivation of the enzyme is a result of addition of a nucleophilic group from the enzyme to the ß carbon of the double bond as indicated by the reaction marked with the X in Scheme III. While there is considerable evidence that this does occur with aspartate 0-decarboxylase (Relyea et al, 1974), we propose that for glutamate decarboxylase the inactivation follows release of -aminoacrylate from its Schiff base by "transimination" (step 2 of Scheme III). Inactivation results from nucleophilic attack by the 0 carbon of the aminoacrylate on the "internal" Schiff base of pyridoxal phosphate with a lysine side chain (step 3 of scheme III).…”

Section: Discussionmentioning

confidence: 88%

“…Many examples are known in which a mechanism-dependent enzyme inhibition occurs when a ß substituent of an inhibitor can be eliminated to form a Schiff base of a-aminoacrylate or a related ,/3-unsaturated amino acid or amine (step 1 of Scheme III; Miles & Meister, 1967;Relyea et al, 1974;John 6 Fasella, 1969; Fowler & John, 1972, 1981; Silverman & Abeles, 1976, 1981Soper et al, 1977;Soper & Manning, 1978;Morino et al, 1974; Wang et al, 1981;Kallio et al, 1981;Bey, 1981). It has generally been assumed, without any real proof, that inactivation of the enzyme is a result of addition of a nucleophilic group from the enzyme to the ß carbon of the double bond as indicated by the reaction marked with the X in Scheme III.…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

A novel reaction of the coenzyme of glutamate decarboxylase with L-serine O-sulfate

Likos¹,

Ueno²,

R³

et al. 1982

Biochemistry

121

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Section: Discussionmentioning

confidence: 88%

Section: Discussionmentioning

confidence: 99%

A novel reaction of the coenzyme of glutamate decarboxylase with L-serine O-sulfate

Likos¹,

Ueno²,

R³

et al. 1982

Biochemistry

121

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“…Pyridoxal S'-phosphate dependent enzymes have often been studied using mechanism-based ("suicide") inhibitors (Badet et al, 1984;Walsh, 1984). For examples, /3-haloalanines have been shown to be effective mechanism-inhibitors of pig heart aspartate aminotransferase (Morino & Okamoto, 1973, alanine aminotransferase (Golichowski & Jenkins, 1978), L-aspartate-/?decarboxylase (Relyea et al, 1974), and serine transhydroxymethylase (Wang et al, 1981). Serine palmitoyltransferase is likely to be affected by such compounds on the basis of its postulated mechanism (Krisnangkura & Sweeley, 1976).…”

mentioning

confidence: 99%

Inhibition of serine palmitoyltransferase in vitro and long-chain base biosynthesis in intact Chinese hamster ovary cells by .beta.-chloroalanine

Medlock¹,

Merrill²

1988

Biochemistry

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The effects of beta-chloroalanine (beta-Cl-alanine) on serine palmitoyltransferase activity and the de novo biosynthesis of sphinganine and sphingenine were investigated in vitro with rat liver microsomes and in vivo with intact Chinese hamster ovary (CHO) cells. The inhibition in vitro was rapid (5 mM beta-Cl-alanine caused complete inactivation in 10 min), irreversible, and concentration and time dependent and apparently involved the active site because inactivation only occurred with beta-Cl-L-alanine (not beta-Cl-D-alanine) and was blocked by L-serine. These are characteristics of mechanism-based ("suicide") inhibition. Serine palmitoyltransferase (SPT) was also inhibited when intact CHO cells were incubated with beta-Cl-alanine (complete inhibition occurred in 15 min with 5 mM), and this treatment inhibited [14C]serine incorporation into long-chain bases by intact cells. The concentration dependence of the loss of SPT activity and of long-chain base synthesis was identical. The effects of beta-Cl-L-alanine appeared to occur with little perturbation of other cell functions: the cells exhibited no loss in cell viability, [14C]serine uptake was not blocked, total lipid biosynthesis from [14C]acetic acid was not decreased (nor was the appearance of radiolabel in cholesterol and phosphatidylcholine), and [3H]thymidine incorporation into DNA was not affected. There appeared to be little effect on protein synthesis based on the incorporation of [3H]leucine, which was only decreased by 14%. Although beta-Cl-L-alanine is known to inhibit other pyridoxal 5'-phosphate dependent enzymes, alanine and aspartate transaminases were not inhibited under these conditions. These results establish the close association between the activity of serine palmitoyltransferase and the cellular rate of long-chain base formation and indicate that beta-Cl-alanine and other mechanism-based inhibitors might be useful to study alterations in cellular long-chain base synthesis.

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“…Initially, the purified protein was assayed in the presence and absence of 3-chloroalanine (3CA). 3CA reacts with PLP in the active site of some PLP-dependent enzymes, where loss of the chlorine substituent generates 2AA which can damage the active site of target enzymes (Badet et al, 1984;Henderson & Johnston, 1976;Relyea et al, 1974). As such, reaction with 3CA can serve as a predictor of sensitivity to free 2AA or the 2IP tautomer.…”

Section: Hem1p Is Damaged By 2aa In Vitromentioning

confidence: 99%

Absence of MMF1 disrupts heme biosynthesis by targeting Hem1pin Saccharomyces cerevisiae

Whitaker

Ernst²,

Downs

2021

Yeast

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The RidA subfamily of the Rid (YjgF/YER057c/UK114) superfamily of proteins is broadly distributed and found in all domains of life. RidA proteins are enamine/imine deaminases. In the organisms that have been investigated, lack of RidA results in accumulation of the reactive enamine species 2-aminoacrylate (2AA) and/or its derivative imine 2-iminopropanoate (2IP). The accumulated enamine/imine species can damage specific pyridoxal phosphate (PLP)-dependent target enzymes. The metabolic imbalance resulting from the damaged enzymes is organism specific and based on metabolic network configuration. Saccharomyces cerevisiae encodes two RidA homologs, one localized to the cytosol and one to the mitochondria. The mitochondrial RidA homolog, Mmf1p, prevents enamine/imine stress and is important for normal growth and maintenance of mitochondrial DNA. Here, we show that Mmf1p is necessary for optimal heme biosynthesis. Biochemical and/or genetic data herein support a model in which accumulation of 2AA and or 2IP, in the absence of Mmf1p, inactivates Hem1p, a mitochondrially located PLP-dependent enzyme required for heme biosynthesis.

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Affinity Labeling of the Active Center of l-Aspartate-β-decarboxylase with β-Chloro-l-alanine

Cited by 31 publications

References 25 publications

A novel reaction of the coenzyme of glutamate decarboxylase with L-serine O-sulfate

A novel reaction of the coenzyme of glutamate decarboxylase with L-serine O-sulfate

Inhibition of serine palmitoyltransferase in vitro and long-chain base biosynthesis in intact Chinese hamster ovary cells by .beta.-chloroalanine

Absence of MMF1 disrupts heme biosynthesis by targeting Hem1pin Saccharomyces cerevisiae

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