l-Phenylalanine ammonia-lyase

Hanson, Kenneth R.; Havir, Evelyn A.

doi:10.1016/0003-9861(70)90100-1

Cited by 143 publications

(65 citation statements)

References 47 publications

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“…3. Although this mechanism includes chemically implausible steps, it has provided the basis for numerous mechanistic investigations for 25 years (1,3,12,(30)(31)(32). While a carbanionic intermediate was postulated, it was not explained how this could be formed.…”

Section: Discussionmentioning

confidence: 99%

“…It has been known for a long time that PAL contains a catalytically essential dehydroalanine residue (3). Although the latter occurs in a number of natural peptides (4, 5) and has been postulated as an intermediate in the conversion of L-serine residues into D-alanine (6), it is shared only by one related enzyme, histidine ammonia-lyase (HAL) (7-10).The electrophilic property of the prosthetic dehydroalanine has been demonstrated by addition of various nucleophiles, such as nitromethane (7-11), cyanide (12), and sodium borohydride (3,8). All these reagents caused inactivation and, using radiolabel, incorporation of the radioisotope into the expected product.…”

mentioning

confidence: 99%

mentioning

confidence: 99%

“…The electrophilic property of the prosthetic dehydroalanine has been demonstrated by addition of various nucleophiles, such as nitromethane (7)(8)(9)(10)(11), cyanide (12), and sodium borohydride (3,8). All these reagents caused inactivation and, using radiolabel, incorporation of the radioisotope into the expected product.…”

mentioning

confidence: 99%

“…All these reagents caused inactivation and, using radiolabel, incorporation of the radioisotope into the expected product. Twenty-five years ago a mechanism was proposed for the PAL and HAL reactions involving as an initial step the nucleophilic attack at the dehydroalanine residue by the amino group of the substrate (3,7). Such a reaction should enhance the leaving ability of the amino group.…”

mentioning

confidence: 99%

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The mechanism of action of phenylalanine ammonia-lyase: the role of prosthetic dehydroalanine.

Schuster

Rétey

1995

Proc. Natl. Acad. Sci. U.S.A.

127

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Phenylalanine ammonia-lyase (EC 4.3.1.5) from parsley is posttranslationally modified by dehydrating its Ser-202 to the catalytically essential dehydroalanine prosthetic group. The codon of Ser-202 was changed to those of alanine and threonine by site-directed mutagenesis. These mutants and the recombinant wild-type enzyme, after treatment with sodium borohydride, were virtually inactive with L-phenylalanine as substrate but catalyzed the deamination of L-4-nitrophenylalanine, which is also a substrate for the wild-type enzyme. Although the mutants reacted about 20 times slower with L-4-nitrophenylalanine than the wild-type enzyme, their V... for L-4-nitrophenylalanine was two orders of magnitude higher than for L-phenylalanine. In contrast to L-tyrosine, which was a poor substrate, DL-3-hydroxyphenylalanine (DL-m-tyrosine) was converted by phenylalanine ammonia-lyase at a rate comparable to that of L-phenylalanine. These results suggest a mechanism in which the crucial step is an electrophilic attack of the prosthetic group at position 2 or 6 of the phenyl group. In the resulting carbenium ion, the 18-Hsi atom is activated in a similar way as it is in the nitro analogue. Subsequent elimination of ammonia, concomitant with restoration of both the aromatic ring and the prosthetic group, completes the catalytic cycle.Phenylalanine ammonia-lyase (PAL) is an important plant enzyme that converts L-phenylalanine into trans-cinnamic acid, which in turn is the precursor of various phenylpropanoids, such as lignins, flavonoids, and coumarins (1, 2). It has been known for a long time that PAL contains a catalytically essential dehydroalanine residue (3). Although the latter occurs in a number of natural peptides (4, 5) and has been postulated as an intermediate in the conversion of L-serine residues into D-alanine (6), it is shared only by one related enzyme, histidine ammonia-lyase (HAL) (7-10).The electrophilic property of the prosthetic dehydroalanine has been demonstrated by addition of various nucleophiles, such as nitromethane (7-11), cyanide (12), and sodium borohydride (3,8). All these reagents caused inactivation and, using radiolabel, incorporation of the radioisotope into the expected product. Twenty-five years ago a mechanism was proposed for the PAL and HAL reactions involving as an initial step the nucleophilic attack at the dehydroalanine residue by the amino group of the substrate (3, 7). Such a reaction should enhance the leaving ability of the amino group. Recently, this laboratory has identified the precursor of the dehydroalanine in HAL from Pseudomonas putida and PAL from parsley to be Ser-143 and Ser-202, respectively (9, 10, 13). Heterologous expression allowed the production of mutants by site-directed mutagenesis. The PAL mutant S202A and the HAL mutants S143A and S143T were virtually inactive with L-phenylalanine and L-histidine as substrates, respectively (9,10,13,14). Surprisingly, L-5-nitrohistidine, known to be a moderately good substrate of HAL, reacted with the dehydroalanine-less m...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 3 more Smart Citations

The mechanism of action of phenylalanine ammonia-lyase: the role of prosthetic dehydroalanine.

Schuster

Rétey

1995

Proc. Natl. Acad. Sci. U.S.A.

127

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Conformational structure of peptides containing dehydroalanine: Formation of ?-bend ribbon structure

Nandel

Malik

Singh

et al. 1999

Int. J. Quant. Chem.

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␣,␤-Unsaturated amino acids dehydroamino acids have been found in naturally occurring antibiotics of microbial origin and in some proteins. Due to the presence of the C C double bond, the dehydroamino acids influence the main-chain between C O of the acetyl group and the NH of the amide group, resulting in the formation of a 10-membered ring. In the model heptapeptide containing ⌬ Ala at alternate position with Ala, Abu, and Leu, the lowest-energy conformation corresponds to s y30Њ and s 120Њ for all the Ala, Abu, and Leu residues and s s 30Њ for all ⌬ Ala residues. A graphical view of the molecule in this conformation reveals the formation of three hydrogen bonds involving the C O moiety of the ith residue and the NH moiety of the i q 3th residue, resulting in a 10-membered ring formation. In this structure, only alternate peptide bonds are involved in the intramolecular hydrogen-bond formation unlike the helices and it has been named the ␤-bend ribbon structure. The helical structures were predicted to be the most stable structures in the heptapeptide Ž . Ac᎐ Aib᎐⌬Ala ᎐NHMe with s "30Њ, s "60Њ for Aib residues and s s "30Њ 3 for ⌬ Ala residues. The computational results reveal that the ⌬ Ala residue does not induce an inverse ␥-turn in the preceding residue. It is the competitive interaction of small solvent molecules with the hydrogen-bonding sites of the peptide which gives rise Ž . to the formation of an inverse ␥-turn s y54Њ, s 82Њ; s 44Њ, s 3Њ in the 1 1 2 2 preceding residue to ⌬ Ala. The computational studies for the positional preference of ⌬ Ala in the peptide containing one ⌬ Ala and nine Ala residues reveals the formation of

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Solid-Phase Synthesis of Peptides Containing α,β-Didehydroamino Acids

et al. 2001

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α,β‐Didehydroamino acids are frequently encountered in natural peptides with important biological activity. Herein, we report a mild and convenient method for the preparation of peptides containing α,β‐didehydroamino acids, where solid‐phase techniques are used both for elongation of the peptide chain and formation of the double bond. This bond is formed through a β‐elimination reaction, using a water soluble carbodiimide as the activating reagent of the hydroxyl function, and catalyzed by CuCl.

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l-Phenylalanine ammonia-lyase

Cited by 143 publications

References 47 publications

The mechanism of action of phenylalanine ammonia-lyase: the role of prosthetic dehydroalanine.

The mechanism of action of phenylalanine ammonia-lyase: the role of prosthetic dehydroalanine.

Conformational structure of peptides containing dehydroalanine: Formation of ?-bend ribbon structure

Solid-Phase Synthesis of Peptides Containing α,β-Didehydroamino Acids

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