Regulation of Bacterial Gene Activity Help (page 3)

By — McGraw-Hill Professional
Updated on Aug 23, 2011

Negative, Repressible Control

An example of a repressible operon under negative control is the tryptophan system of E. coli (Fig. 10-12).

Negative, Repressible Control

The amino acid tryptophan is synthesized in five steps, each step mediated by a specific enzyme. The genes responsible for these five enzymes are arranged in the trp operon in the same order as their enzymatic protein products function in the biosynthetic pathway. The regulatory gene for this system constitutively synthesizes a nonfunctional protein called aporepressor. When tryptophan is in oversupply, the excess tryptophan acts as a corepressor by binding to the aporepressor forming a functional repressor complex. The functional repressor binds to the trp operator and coordinately represses transcription of all five structural genes in the operon. The promoter and operator regions overlap significantly, and binding of active repressor and RNA polymerase are thus competitive. When tryptophan is in lowconcentration, tryptophan dissociates from the aporepressor, and the aporepressor protein falls off the operator. RNA polymerase then synthesizes the polycistronic mRNA for all five enzymes of the tryptophan pathway.

A secondary regulatory mechanism also exists in the tryptophan operon. At the 50 end of the polycistronic mRNA of this operon a 162 base pair leader sequence precedes the coding segments for the five enzymes. There are two adjacent tryptophan codons in the leader peptide. When tryptophan is present in excess, transcription of the rest of the trp operon is prevented because RNA polymerase generates a transcription termination sequence; this phenomenon is known as attenuation. A model (Fig. 10-13) that explains attenuation assumes that (when tryptophan is abundant) movement of bacterial ribosomes follows closely behind the movement of RNA polymerase as it synthesizes mRNA, and all intra molecular base pairing is prevented in the mRNA segment in contact with the ribosome. In the experimental absence of ribosomes, only the leader mRNA is transcribed and no translation occurs [Fig. 10-13(a)]. Leader segments 1 and 2 become folded into stem and loop A by complementary base pairing, whereas segments 3 and 4 fold into stem and loop C that acts as a transcription termination signal. As RNA polymerase synthesizes the seven uracils that follow segment 4, these uracils and the adjacent paired 3-4 region of mRNA (having just folded into stem and loop C) form a terminator signal that causes RNA polymerase to prematurely dissociate from the DNA before it can transcribe any of the DNA coding segments for the five enzymes of the trp operon.

Negative, Repressible Control

When only the concentration of activated trp-tRNAs is low[Fig. 10-13(b)] ribosomes begin to translate region 1, thereby preventing pairing of regions 1 and 2. However, the ribosome tends to stall momentarily (especially at the pair of tryptophan codons), allowing pairing of regions 2 and 3 to forma B stem-and-loop structure (called an antiterminator); regions 3 and 4 are thereby prevented from forming the C termination signal, and RNA polymerase is allowed to continue transcription on into the trp operon.

If activated trp-tRNAs are abundant [Fig. 10-13(c)], ribosomes follow so closely behind RNA polymerase that the anti-terminator B structure cannot form, and therefore the terminator C structure does form. Thus, only the leader peptide (but none of the five enzymes of the operon) can be translated from the prematurely terminated mRNA.

The repressor mechanism coarsely regulates the tryptophan system, whereas the attenuation mechanism fine tunes the control over tryptophan concentrations. Attenuation of the trp operon is also sensitive to the concentrations of several amino acids other than tryptophan. Operons for the amino acids histidine and leucine, however, are thought to be regulated only by attenuation.

Positive, Inducible Control

An example of a positive, inducible regulatory mechanism is found in the arabinose operon of E. coli. Arabinose is a sugar that requires three enzymes (coded by genes araB, araA, araD) for its metabolism. Two additional genes are needed to transport arabinose across the cell membrane, but they are located at a distance from the BAD cluster coding for the catabolic enzymes. The regulatory gene araC is close to the promoter for the BAD cluster. The protein product of the araC gene (AraC) is a repressor of the BAD cluster when the substrate arabinose is absent. However, when arabinose is present, it binds to the repressor (AraC), forming an activator complex that facilitates the binding ofRNApolymerase to the promoter, thus inducing transcription of the operon. The preceding story is a gross oversimplification of the complexity that is already known about the regulation of the arabinose system. For example, cyclic adenosine monophosphate (cAMP) and catabolite activator protein (CAP; also known as cyclic AMP receptor protein, CRP) are also involved in the regulation of the arabinose system. The action of these last two molecules in the phenomenon of catabolite repression is discussed in the next section.

Global Regulation or Multiple Controls

A genetic locus may be regulated by more than one mechanism. When glucose is available, there is no need to catabolize other sugars, and the genes coding for these other sugar-catabolizing enzymes can be turned off. For example, if glucose is absent and lactose is present in the medium, the lac operon would be induced. But if glucose is present, induction of the lac operon does not occur. This phenomenon was originally termed the glucose effect; it is now known as catabolite repression. A complex of two molecules, namely, cAMP and CAP, acts as the activator in catabolite repression.Within the lac promoter (Fig. 10-11), there is a site for binding of a cAMP-CAPcomplex.RNA polymerase only binds effectively to the promoter if cAMP-CAP complex is also bound to this site. As the level of glucose increases within the cell, the amount of cAMP decreases and less cAMP-CAP complex is available to activate the lac operon. CAP is produced at low levels by its own genetic locus. The enzyme adenylate cyclase (adenylcyclase) converts adenosine triphosphate (ATP) to cyclic adenosine monophosphate (cAMP). Adenylate cyclase can become activated to first messenger status by the interaction of specific cell receptors with their target molecules; the cAMP thus produced (second messenger) can then regulate a battery of genes coordinately.

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