The trp operon is a genetic unit found in bacteria, such as E. coli, that regulates the cell’s supply of the amino acid Tryptophan. An operon is a set of linked genes and their regulatory sequences transcribed together under the control of a single promoter, allowing for coordinated gene expression in prokaryotes. The trp operon ensures the cell produces the necessary enzymes for Tryptophan biosynthesis only when the amino acid is scarce in the surrounding environment. By controlling these production genes, the cell conserves energy and resources by not synthesizing a compound it can acquire readily from its surroundings.
The Components and Goal of the trp Operon
The trp operon includes several distinct DNA segments necessary for its function. Five structural genes, designated \(trpE, trpD, trpC, trpB,\) and \(trpA\), encode the enzymes required to convert a precursor molecule into Tryptophan. These five genes are transcribed into a single messenger RNA molecule.
Upstream of these structural genes are the regulatory components that dictate whether transcription proceeds. The promoter sequence serves as the binding site for the RNA polymerase enzyme, which is responsible for initiating transcription. Adjacent to the promoter lies the operator, a short DNA segment where a regulatory protein can bind to physically block the movement of RNA polymerase.
The operon also includes a region called the leader sequence, located between the operator and the first structural gene, \(trpE\). The overall biological goal of this entire assembly is to precisely match the cell’s production rate of Tryptophan with its immediate metabolic need. This system allows the bacterium to rapidly adjust its enzyme production, thereby avoiding the energetic cost of synthesizing an amino acid that is already available.
Regulation by the trp Repressor
The primary control of the operon involves a molecule called the trp repressor protein. This repressor is encoded by a separate gene, \(trpR\), which is located elsewhere on the bacterial chromosome and is expressed continuously at a low level. The repressor protein exists in an inactive form that cannot bind to the operon’s DNA.
When the concentration of Tryptophan within the cell rises, the amino acid acts as a corepressor by binding directly to the inactive repressor protein. This binding causes a change in the repressor’s three-dimensional shape, converting it into its active form. The newly activated repressor protein then has a high affinity for the operator region of the trp operon.
Binding of the Tryptophan-repressor complex to the operator physically obstructs the path of RNA polymerase, preventing the enzyme from transcribing the structural genes. This mechanism effectively shuts down Tryptophan synthesis. This type of control is known as a negative repressible system.
Conversely, when Tryptophan levels drop, the amino acid dissociates from the repressor protein, causing the repressor to revert to its inactive shape. Because the inactive repressor cannot bind to the operator, RNA polymerase is free to initiate transcription of the structural genes. This primary mechanism alone can reduce transcription of the operon by approximately 70-fold.
Fine-Tuning Control: Attenuation
Attenuation provides a secondary, fine-tuning adjustment to gene expression that operates after transcription has already begun. This mechanism relies on the coupling of transcription and translation, which is possible in prokaryotes because they lack a nucleus. The key to attenuation is the leader sequence, which is transcribed into a region of mRNA, known as the leader transcript, before the structural genes.
This leader transcript contains four distinct sequences, numbered 1, 2, 3, and 4, each of which is partially complementary to the next. These complementary regions allow the mRNA to fold into various stem-loop or “hairpin” secondary structures, which dictate the fate of the ongoing transcription process. A small open reading frame within region 1 encodes a short leader peptide and crucially contains two adjacent Tryptophan codons.
The ribosome acts as the sensor in this system, tracking the availability of Tryptophan-charged transfer RNA molecules (\(tRNA^{Trp}\)) as it attempts to translate the leader peptide.
High Tryptophan Levels
When Tryptophan levels are high, \(tRNA^{Trp}\) is readily available, allowing the ribosome to translate the two Tryptophan codons quickly without pausing. The fast-moving ribosome reaches the boundary between regions 1 and 2, physically covering region 2 before region 3 is completely transcribed.
Because region 2 is blocked, the newly synthesized region 3 is forced to pair with region 4, forming a 3-4 hairpin structure. This 3-4 structure is a transcriptional terminator, often followed by a sequence of uracil residues, which signals the RNA polymerase to dissociate from the DNA. Transcription is prematurely terminated, resulting in a short, non-functional mRNA and a high degree of repression.
Low Tryptophan Levels
Conversely, when Tryptophan levels are low, the cell lacks sufficient \(tRNA^{Trp}\), and the ribosome stalls immediately upon encountering the two Tryptophan codons in region 1. This stalling causes the ribosome to physically shield region 1, which prevents region 1 from pairing with region 2. With region 2 now available, it immediately pairs with the newly transcribed region 3, forming a 2-3 hairpin structure.
The 2-3 hairpin is known as the anti-terminator structure because its formation prevents the creation of the 3-4 terminator hairpin. With the terminator structure absent, RNA polymerase continues transcribing past the leader sequence and into the structural genes, allowing the cell to synthesize the enzymes needed to make its own Tryptophan. This attenuation mechanism provides a graded response, acting like a dimmer switch to adjust Tryptophan production based on minute changes in amino acid availability, further reducing transcription by another factor of ten beyond the repressor’s effect.