Derepression of enzyme synthesis

Figure 11-1 Some control elements for metabolic reactions. Throughout the book modulation of the activity of an enzyme by allosteric effectors or of transcription and translation of genes is indicated by dotted lines from the appropriate metabolite. The lines terminate in a minus sign for inhibition or repression and in a plus sign for activation or derepression. Circles indicate direct effects on enzymes, while boxes indicate repression or induction of enzyme synthesis.

Synthesis of many enzymes is repressed most of the time. The appearance of an enzyme at a particular stage in the life of an organism as well as the differing distributions of isoenzymes within differentiated tissue result from derepression. The control of enzyme synthesis may also be exerted during the splicing of transcripts and at the translational level as well. These control mechanisms are often relatively slow, with response times of hours or even days. However, effects on the synthesis of some hormones, such as insulin (Section G), may be observed within a few minutes. 

Candida utilis has high levels of NAD-GDH when grown on either glutamate or some other amino acids. Addition of either ammonia or glutamine to yeast so adapted can result in rapid and extensive inactivation of NAD-GDH at a faster rate than can be accounted for by the immediate cessation of enzyme synthesis (331). The NADP-GDH is not subject to rapid changes in activity under these conditions. In Saccharomyces carlsbergensis synthesis of NAD-GDH is also repressed by NH4+ and derepressed by glutamate the reverse is true for NADP-GDH (90,238). 

Induction (derepression) and repression of enzyme synthesis by transcriptional regulation on the DNA template, or the RNA polymerase. 

Fig. 3 Formation of acetylornithinase in Escherichia coli under various regulatory conditions. Plots I and 2 represent formation of the enzyme in the wild-type strain W cultivated without or with added L-arginine hydrochloride (0.2 mg/ml), respectively. Plot 3 represents the formation of the enzyme in the argR mutant W2D (which has an incidental pro marker and is grown in the presence of L-proline, 0.1 mg/ml). Enzyme and total-protein concentrations are expressed per milliliter of extract (1 ml corresponding to 7.5 ml of original culture). The slopes of the plots are the so-called diflFerential rates of enzyme synthesis indicating partial repression (7), full repression (2), and genetic derepression (i).

Further evidence for the involvement of finished enzymes has come from an examination of the rates of enzyme synthesis in the arginine (Lavalle [77]) or tryptophan (Lavalle and De Hauwer [78]) pathways of E. coli K12, on addition of arginine or tryptophan, respectively, to physiologically derepressed cultures There are prompt and sharp... 

The factors that determine the expression of one pathway over another are not known. Activation or inactivation of enzymes may result from derepression or repression of the genome or modification of the rate of enzyme synthesis by regulation of transcription or by regulation of positive or negative feedback mechanisms. 

As expected, in vitro transcription assays involving PARP-1, NAD, and PARC illustrate these predicted outcomes (Kim et al, 2004). Even when driven by a transcriptional activator, such as estradiol-bound estrogen receptor, transcription is repressed when PARP-1 is added to chromatin templates. The repression is reversed by NAD+, and the NAD+-dependent effects are reversed by PARC (Kim et al, 2004). This system for transcriptional control shifts new importance onto the enzymes responsible for synthesis of NAD+ in the nucleus, such as nicotinamide mononucleotide adenylyltransferase-1 (Magni et al, 2004). Because NAD+ facilitates the decompaction of chromatin and the derepression of transcription, nuclear NAD+ biosynthetic enzymes may play critical roles as cofactors. 

As well as induction of the synthesis of the apoprotein portion of cytochrome P-450, there is also induction of the synthesis of the heme portion. Clearly, it is also necessary to have an increased amount of heme if there is an increase in the amount of the enzyme apoprotein being synthesized. Thus, the rate-limiting step in heme synthesis, the enzyme 5-aminolaevulinate synthetase, is inducible by both phenobarbital and TCDD. This is the result of transcriptional activation of the gene, which codes for the S-aminolaevulinate synthetase. It may be that the decrease in the heme pool, which results from incorporation of heme into the newly synthesized apoprotein, leads to derepression of the gene and hence increased mRNA synthesis. The gene repression could be heme-mediated, or heme may modulate P-450 genes. 

There are basically three types of cytochrome P450 inducers (1) phenobarbital-like (the major class) (2) methylcholanthrene-like (which actually increases a P448 isozyme) and (3) anabolic steroids. The former two have been the most frequently studied. Research over the past 40-50 years indicates that their mechanism of action involves genetic interaction, possibly via derepression of a repressor gene, and the subsequent synthesis of mRNA for the specific enzyme proteins. Examples of phenobarbital-like enzyme inducers, the most common, are shown in Table 3.6. 

The exbB mutants (47) are derepressed in enterobactin synthesis and produce this siderophore in iron-containing media. The means whereby iron represses enterobactin synthesis is still obscure. Several years ago it was noted that growth of E. coli on low iron media led to changes in various fRNAs (77). In E. coli K-12 aromatic amino-acid synthesizing enzymes are also derepressed in low iron media, possibly because of the diversion of the chorismate pool to enterobactin (78). 

Cyclic AMP can affect the synthesis of catecholamines by two separate modes of action on the rate-limiting catecholamine-synthesizing enzyme tyrosine hydroxylase. Cyclic AMP-dependent kinase can phosphorylate tyrosine hydroxylase, which in turn results in activation of the enzyme. Furthermore, in peripheral tissues such as the adrenal medulla, cAMP can result in the de novo synthesis of tyrosine hydroxylase by causing a derepression of gene expression due to the translocation of the catalytic subunit of the kinase to the chromaffin cell nucleus. 

Analogs and Inhibitors, When we use prototrophic microorganisms, production of repressors can be reduced by addition of end product analogs to the medium. Thus, all 10 histidine pathway enzymes are derepressed up to 30-fold by 2-thiazolealanine (Ames and Hartman, 1963). In a similar manner, adenine, which inhibits thiamine synthesis, derepresses the enzymes of thiamine biosynthesis (Kawasaki et al., 1969). 

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