Chorismate mutase isozymes

Table II. Differential Properties of Chorismate Mutase Isozymes...

To what extent is the response of cytosolic and plastidic isozymes of the shikimate pathway coordinated or coupled with one another and to alterations in expression of enzymes of the flavonoid and phenylpropanoid-pathway segments Some of the emerging information is given in Figure 6. Thus, light induction, well known to induce PAL and enzymes of the flavonoid pathway, also induces both DS-Mn and DS-Co in parsley cell cultures (49). However, only the cytosolic CM-2 (and not the plastidic CM-1) was induced. Fungal elicitor was reported to induce only DS-Mn—not DS-Co or either of the chorismate mutase isozymes (49). Previous studies... 

Figure 6. Effects of various treatments or manipulations upon levels of phenylalanine ammonia-lyase (PAL), chalcone synthase (CHS), and the separately compartmented isozymes of DAHP synthase and chorismate mu-tase. Upwardly pointed arrows indicate a positive response (enzyme elevation) to the indicated treatment, whereas horizontal arrows indicate no change in enzyme level. References documenting the results shown are line 1 (54 our results with DAHP synthase and chorismate mutase isozymes) line 2 (49,55) line 3 (49,50,56) line 4 (57) line 5 (51) line 6 (52).

In higher plants, phenylalanine seems to be formed in an alternative manner by formation of prephenate (26) and conversion of this intermediate into arogenic acid (32). Chorismate mutase occurs as two isozymes which have been purified to homogeneity from mung bean and sorghum. All chorismate mutase isozymes show allosteric activation by chorismate (Poulsen and Verpoorte, 1991). One chorismate mutase isozyme is inhibited by phenylalanine or tyrosine and activated by tryprophan, whereas the second is not affected by any of the aromatic amino acids. 

If stationary-phase physiology of cultured-cell populations is equated with the slowed growth of mature organismal tissues, then the results obtained with the chorismate mutase isozyme pair (Fig. 3) seem to be consistent with the results shown in Figure 5 for the isozymes of DAHP synthase. Thus, cytosolic CM-2 is elevated in organismal tissue and cytosolic DAHP synthase-Co (DS-Co) is elevated during stationary-phase physiology of tissue culture. 

Figure 1. Hypothetical mechanism for shuttling of intermediates of the common aromatic pathway between plastidic and cytosolic compartments. Enzymes denoted with an asterisk (DAHP synthase-Co, chorismate mutase-2, and cytosolic anthranilate synthase) have been demonstrated to be isozymes located in the cytosol. DAHP molecules from the cytosol are shown to be shuttled into the plastid compartment in exchange for EPSP molecules synthesized within the plastid. Abbreviations C3, phosphoenolpyruvate C4, erythrose 4-P DAHP, 3-deoxy-D-arabino-heptulosonate 7-phosphate EPSP, 5-enolpyruvylshikimate 3-phosphate CHA, chorismate ANT, anthranilate TRP, L-tryptophan PPA, prephenate AGN, L-arogenate TYR, L-tyrosine and PHE, L-phenylalanine.

Of the separately compartmented isozyme pairs that exist for DAHP synthase, chorismate mutase, and anthranilate synthase, each isozyme member of a given pair has different properties of regulation and other distinctive characteristics (see Tables I and II). This suggests a high probability that each isozyme is the gene product of a different gene. 

We have examined the time course of changes induced in isozymes of chorismate mutase and DAHP synthase in potato tubers following mechanical wounding (Table III). In each case both isozymes responded—the plastidic isozyme responding sooner and to a greater extent than the cytosolic isozyme. All five of the other pathway enzymes so far examined were induced by mechanical wounding. 

Two distinct isozymes of chorismate mutase have been demonstrated in N. silvestris. Form CM-1 is subject to allosteric control by phenylalanine, tyrosine and... 

The CM-1 and CM-2 isozymes of chorismate mutase in N. silvestris have been shown to exist in plastidial and cytosolic compartments. Similar evidence also exists in support of a parallel compartmentation of DAHP synthase isozymes, DAHP synthase-Mn being plastid-localized and DAHP synthase-Co being cytosolic (d Amato and Ganson, unpublished data), Prephenate aminotransferase activity of N. silvestris is also largely or entirely localized within plastids (d Amato and Bonner, unpublished data). 

In one experiment washed chloroplasts were isolated and assayed for nitrite reductase, DAHP synthase-Mn and chorismate mutase-1 activities (Table 3). Since enzymes may fractionate with organelles by non-specific (or specific) association with the organelle surface, latency determinations were made. With this approach, activity determinations are made before and after rupture of the washed chloroplasts. If activities are located within the organelle, they are expected to increase dramatically following organelle disruption. Thus, nitrite reductase (chloroplast marker enzyme) gave a latency value of 16, a value similar to those obtained for DAHP synthase-Mn and chorismate mutase-1. The identity of chorismate mutase as the CM-1 isozyme was confirmed by its sensitivity to inhibition by L-tyrosine. 

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