Phenylalanine ammonia-lyase inhibitors

Phenylalanine ammonia-lyase (PAL EC 4.3.1.5) is a pivotal enzyme in controlling flow of carbon from aromatic amino acids to secondary aromatic compounds (Figure 1) (28). PAL primarily deaminates phenylalanine to form t-cinnamic acid, however, in many species, it also less efficiently deaminates tyrosine to form -coumaric acid. Because PAL is restricted to plants and is an important enzyme in plant development, Jangaard (29) suggested that PAL inhibitors might make safe and effective herbicides, however, in his screen of several herbicides, he found no compound to have a specific effect on PAL. This was also the case in studies by Hoagland and Duke (30, 31.) in which 16 herbicides were screened. 

Inhibition of synthesis of the aromatic matrix by inhibitors of phenylalanine ammonia lyase causes the inhibition of deposition of aliphatic components and prevents development of diffusion resistance. Inhibition of synthesis of peroxidase, the enzyme involved in the deposition of the polymeric phenolic matrix, caused by iron deficiency, prevents deposition of aliphatic components of suberin. 

J Zoh, N Amrhein, R Gancarz (2002) Inhibitors of phenylalanine ammonia-lyase 1-aminobenzylphosphonic acid substituted in the benzene ring Phytochemistry 59(1) 9-21... 

Zont, J. and Amrhein, N. (1992) Inhibitors of phenylalanine ammonia-lyase 2-aminoindan-2-phosphonic acid and related compormds. Liebigs Ann. Chem., 625-8. 

Amrhein, N., K.H. Godeke, and V.I. Kefeli (1975). The estimation of relative intracellular phenylalanine ammonia-lyase(PAL)-activities and the modulation in vivo and in vitro by competitive inhibitors. Ber. Deutsch.Bot.Ges.. 89, 247-259. 

The beginning of secondary product formation is often directly coupled with the synthesis of the corresponding enzymes. One example is the biosynthesis of flavonoids in cell cultures of Petroselinum hortense (Fig. 7). Here the regulatory mechanisms were extensively investigated with phenylalanine ammonia-lyase (PAL) (D 22.2.1) and chalcone synthase (D 22.3.3), the key enzymes of the biosynthetic chain. Experiments with inhibitors of transcription and translation showed that the increase of enzyme activity depends on RNA and protein biosynthesis. Labeling experiments demonstrated that it is caused by an accelerated rate of enzyme synthesis. 

In this chapter, the discussion will concentrate on two inhibitors with a reasonable claim to selective action on enz3ones related to the shikimate pathway glyphosate, which inhibits 5-enolpyruvylshikimate 3-phosphate (EPSP) synthase and L-a-aminooxy-3 phenylpropionic acid (L-AOPP), an inhibitor of phenylalanine ammonia-lyase (PAL) (Fig. 2). In addition to introducing a novel inhibitor of PAL, (R)-(l-amino-2-phenylethyl)phosphonic acid (APEP), previous and current efforts to design inhibitors of other shikimate pathway enzymes will be described. The treatment presented here will show that the deductions and predictions made on the basis of the abstract scheme in Figure 1 can be, and have been, tested on the basis of the real pathway presented in Figure 2. 

While the results of the growth reversal experiments in the "supercomplex systems" described above had stimulated investigations at the level of "defined systems" (i.e. the examination of certain metabolic reactions in cell-free systems), no definitive answers as to the biochemical mode of action of glyphosate had been obtained. Based on our experience with inhibitors of phenylalanine ammonia-lyase (see below), we included the "complex system" level (i.e. the examination of a metabolic pathway jsi vivo) in our strategy in order to define the limits more closely. Hypocotyls from etiolated buckwheat seedlings provided a system in which the rapid synthesis of phenylalanine-derived products, such as anthocyanin and other phenylpropanoid compounds, can be very simply induced by illumination. Anthocyanins, in particular, can be conveniently extracted and quantified and, at least in buckwheat, are not subject to measurable turnover within... 

Phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) is not considered an enzyme of the shikimate pathway proper, but since in higher plants it channels a large flux of fixed carbon into phenolic compounds, in particular lignin, the shikimate pathway must furnish its substrate, phenylalanine, in sufficient amounts. Literature on PAL has periodically been reviewed (literature in References 88 and 89) and I will concentrate here on a selective and brief discussion of our work on PAL inhibitors in the Bochum laboratory. 

Table 2. Inhibition constants (Ki) of competitive inhibitors of buckwheat phenylalanine ammonia-lyase...

Inhibition of lignin formation by L-a-aminooxy-8-phenylpropionic acid, an inhibitor of phenylalanine ammonia-lyase. Eur. J. Cell Biol. 29 139-144. 

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