Monday, October 17, 2011

The Trp Operon

The last post about an operon (the lac operon) is the most viewed post on this blog, so I thought that it might be helpful to follow this up with another operon, this time concentrating on the trp (tryptophan) operon.  This operon is another really elegant example of transcriptional regulation in E.coli and the mechanism is pretty cool.

Amino acids are essential for life (see the last post on their composition!) and cells synthesize amino acids using a variety of enzymes.  When nutrients are plentiful, such as E.coli would encounter in nutrient broth in the laboratory setting, cells no longer need to waste energy producing biosynthetic enzymes when they can utilize nutrients already in excess.  The trp operon contains several enzymes that are coordinately regulated and involved in the production of tryptophan.  When tryptophan is present in the cell's environment, it doesn't need to make any of these enzymes, but if the cell needs tryptophan, these enzymes are transcribed and shortly thereafter translated.  Control of this operon, thus, controls how much energy the cell is going to put into making tryptophan.

Similar to the lac operon, the trp operon contains an operator (O) sequence, within the promoter sequence, where an operator binds and prevents transcription.  In the presence of tryptophan, the operator binds the promoter and prevents RNA polymerase from transcribing genes.  In the absence of tryptophan, however, transcription occurs at a basal rate.  Sounds simple enough, right?  Let's take it a step further and consider...

An important concept in gene regulation is that of attenuation, which is fine-tuning of gene expression.  You might think that attenuation is mediated by protein factors that bind the DNA and affect gene expression; however, attenuation of the trp operon is a little different and, instead, depends on mRNA structure to modulate gene expression.

Before moving forward, let's look at the trp operon (diagrammed to the right).  Briefly,t here are four regions, and these four regions have differing levels of complementarity to each other.  Thus, when the DNA is transcribed into mRNA, the mRNA folds into all kinds of shapes and the regions of the trp operon fold on each other.

After transcription of the entire trp operon (we're dealing with mRNA from this point forward), the next event is translation of this mRNA into protein.  In bacteria, it's important to note that transcription and translation occur simultaneously, so as soon as we have a transcript in a bacterial cell, it's being translated.  The trp transcript contains two critical tryptophan codons immediately before region 1, so in order to synthesize the enzymatic machinery to make tryptophan, the cell must use a few residues to translate the protein.

In the presence of high amounts of tryptophan within the cell, the ribosome plows through these two tryptophan codons, adding in the appropriate amino acids, and continuing through region 1 of the mRNA.  This results in region 1 and 2 mRNA sequences binding together, and then regions 3 and 4 bind together as well.  This interaction between regions 3 and 4 results in the creation of a transcription-termination hairpin, basically a structure in the mRNA that kicks out RNA polymerase and prevents further transcription of the mRNA.  Thus, transcription (and then translation) are stopped because

In the absence of tryptophan, however, the ribosome cannot quickly add tryptophan during the translation process and it stalls before region 1.  This results in the folding of the mRNA such that regions 2 and 3 bind to each other.  When this structure forms, no transcriptional termination hairpin is formed, and mRNA synthesis continues.  Thus, the entire mRNA sequence for the trp operon is made and can be translated into enzymes that will synthesize tryptophan.

In summary:
Lots of tryptophan: Ribosome zooms through the mRNA, regions 1 & 2 and 3 & 4 bind (in pairs) and create a termination hairpin
End result: Transcription terminates and tryptophan synthetic enzymes not created (cell saves energy!)

Lack of tryptophan: Ribosome stalls immediately before region 1, regions 2 and 3 bind each other, no termination hairpin is formed
End result: Transcription continues and biosynthetic enzymes are eventually synthesized

This scheme is similar for other operons encoding amino acid biosynthetic enzymes (in bacteria, that is).  The trp operon is an elegant scheme to finely-tune transcription via mRNA structure to prevent the cell from wasting energy.

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