Tyrosine phenol lyases

Inhibition of enzymes by fluorinated derivatives of tyrosine have been the focus of many investigations. The inhibition of tyrosine phenol lyase,of... 

The three-dimensional and primary structures of the closely related enzyme, tyrosine phenol-lyase, have recently been determined.28 Since 43% of the amino acid sequences of this enzyme and tryptophanase is identical, their three-dimensional structures should be very similar. The structure of the lyase is like a butterfly. The tetrameric molecule can be considered from a crystallographic point of view as an a2a 2-structure consisting of two... 

There is at the moment no compelling evidence for either of these mechanisms. An important experiment which needs to be done with enzymes of this class is to probe for internal transfer of the a-hydrogen from one enantiomer to the other under single turnover conditions with trapping of the product. An experimental design to accomplish this is currently being explored with tyrosine phenol-lyase and will be discussed below. Demonstration of any internal return of the a-hydrogen... 

The closely related bacterial enzyme tyrosine phenol-lyase [137] has an even wider substrate and reaction specificity than tryptophanase, including the remarkable ability to cleave both D- and L-tyrosine and to interconvert D- and L-alanine. As already discussed and summarized in Tables 1 and 2, the stereochemistry at C-/J in all the a,/3-elimination and -replacement reactions of this enzyme studied so far is always retention [108,109,129]. This includes the a, -elimination of L- as well as of D-tyrosine. The fate of the a-hydrogen of L-tyrosine in this reaction has been probed in preliminary experiments (H. Kumagai, E. Schleicher and H.G. Floss, unpublished results), and the results tentatively suggest transfer of deuterium from the a-position to C-4 of the resulting phenol. Attempts to demonstrate intramolecularity of this transfer have so far been inconclusive. The base abstracting H-a in this enzyme may be histidine [138]. 

A system has recently been developed to examine the question whether any internal return of the H-a can be demonstrated in the interconversion of L- and D-alanine catalyzed by this enzyme. L-[2-2H]Alanine in HzO or unlabeled L-alanine in 2H20 are converted into D-alanine with tyrosine phenol-lyase in the presence of excess D-amino acidiacetyl-coenzyme A acetyl transferase [139] and acetyl-coenzyme A to ensure single turnover conditions. The resulting TV-acetyl-D-alanine is analyzed for its deuterium content by mass spectrometry and NMR. Initial data indicate 12% transfer of a- H from L-alanine to the D-isomer in 2HzO (S.-j. Shen, H. Kumagai and H.G. Floss, unpublished results), but these results need to be verified. If they are confirmed, racemization by a single base mechanism would be indicated. 

Another example of how catalysis plays a key role in enabling our lives is in the synthesis of pharmaceuticals. Knowles s development, at Monsanto in the early 1970s, of the enantioselective hydrogenation of the enamide precursor to L-DOPA (used to treat Parkinson s disease), using a Rh-chiral phosphine catalyst (Section 3.5), led to a share in the Nobel prize. His colaureates, Noyori and Sharpless, have done much to inspire new methods in chiral synthesis based on metal catalysis. Indeed, the dramatic rise in the demand for chiral pharmaceutical products also fuelled an intense interest in alternative methodologies, which led to a new one-pot, enzymatic route to L-DOPA, using a tyrosine phenol lyase, that has been commercialized by Ajinomoto. 

Tyrosine phenol lyase (p-tyrosinase) has been shown to catalyze the efficient synthesis of the L-amino acids L-tyrosine and L-dopa from pyruvate, ammonia and phenol, or catechol, respectively (87-89). 

D- and L- Serine dehydratases (deaminases) Tryptophan indole-lyase (tryptophanase) Tyrosine phenol-lyase ... 

L-Aspartate L-DOPA Fumarate Pyruvate, catechol, NH4 Aspartase Tyrosine-phenol lyase 6000 tons 200 tons... 

The structures and properties of quinone methides were recently reviewed . Inter alia, the microbial tyrosine phenol lyase (TPL) catalyzes the a, /3-elimination of L-tyrosine to phenol and ammonium pyruvate. It is assumed that the process includes three steps, the second of which is tautomerization of the aromatic moiety which converts it into a good leaving group (equation 49) °. 

L-Dopa Addition of ammonia with L-tyrosine phenol lyase from Erwinia herbicolad 250 t/y Ajinomoto [9]... 

A further industrially important lyase for the production of L-amino acids is the tyrosine phenol lyase [39]. This biocatalyst is used by Ajinomoto in the production of the pharmaceutically important L-3,4-dihydroxyphenylalanine (L-dopa), 32, which is applied in the treatment of Parkinson s disease. The reaction concept is based on a one-pot three-component synthesis starting from catechol, 30, pyruvic acid, 31, and ammonia in the presence of suspended whole cells (strain Erwinia herbicola) containing the tyrosine phenol lyase biocatalyst (Fig. 16). A key feature of this process is the high volumetric productivity of 110 g/L of the desired L-dopa product. Notably, this reaction runs with an annual capacity of 250 tons. 

Fig. 16 Production of L-dopa using tyrosine phenol lyase.

Our laboratory has studied the stereochemistry of methyl group formation in a number of a, 0 elimination reactions of amino acids catalyzed by pyridoxal phosphate enzymes. The reactions include the conversions of L-serine to pyruvate with tryptophan synthase 02 protein (78) and tryptophanase (79), of L-serine and l-tyrosine with tyrosine phenol-lyase (80), and l-cystine with S-alkylcysteine lyase (81). In the latter study, the stereospecific isotopically labeled L-cystines were obtained enzymatically by incubation of L-serines appropriately labeled in the 3-position with the enzyme O-acetyl serine sulfhy-drase (82). The serines tritiated in the 3-position were prepared enzymatically starting from [l-3H]glucose and [l-3H]mannose by a sequence of reactions of known stereochemistry (81). The cysteines were then incubated with 5-alkyl-cysteine lyase in 2H20 as outlined in Scheme 19. The pyruvate was trapped as lactate, which was oxidized with K2Cr202 to acetate for analysis. Similarly, Cheung and Walsh (71) examined the conversion of D-serine to pyruvate with... 

P. Christen, Conversion of tyrosine phenol-lyase to dicarboxylic amino acid P-lyase, an enzyme not found in nature, J. Biol. Chem. 1999, 274(3), 1320-1325. 

Figure 14.6-1. Synthesis of p-substituted a-amino acids from p-chloroalanine using tryptophan synthase and tyrosine phenol lyase.

Synthesis of L-Dopa Catalyzed by Tyrosine Phenol Lyase from Erwinia herbicola (E.C. 4.1.99.2)1100-1031... 

E = tyrosine phenol lyase, whole cells from Erwfnia herbicola... 

Tyrosine-phenol lyase/L-DOPAa Enterobacteriaceae see text... 

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