TRYPTOPHAN OPERON -FOR CSIR NET

            

TRYPTOPHAN OPERON:

         Tryptophan – α-amino acid used in the biosynthesis of proteins.

            Operon  -   Functional unit of DNA containing cluster of gene under a single promoter.

structure of tryptopha

                                      

ENZYMES NEEDED FOR BIOSYNTHESIS AND THE GENES WHICH ENCODE THEM:

GENES	ENZYMES
trp E	Anthranilate synthase
trp D	Anthranilate phosphoribosyl transferase
trp C	Phosphoribosyl anthranilate isomerase, indole 3-glycerol phosphate synthase
trp B	Tryptophan synthase (B protein)
trp A	Tryptophan synthase (A protein)

 

·       The tryptophan (trp) operon of E. coli consists of five structural  genes that encode enzymes needed for synthesis of the amino acid tryptophan.

·        It is regulated by the trp repressor,which is encoded by the trpR gene. Because the enzymes encoded by the trp operon function in a biosynthetic pathway, it is  wasteful to make the enzymes needed for tryptophan synthesis when tryptophan is readily available.

low level of trp -so trancription of entire trp operon occurs

              

·       Therefore the operon functions only when tryptophan is not present and must be made de novo from precursor molecules.

·        To accomplish this regulatory goal, the trp repressor is synthesized in an inactive form that cannot bind the trp operator as long as tryptophan levels are low.

high level of trp - so repression occurs

                  

·       When tryptophan levels increase, tryptophan acts as a co-repressor, binding the repressor and activating it.

·       The repressor-corepressor complex then attaches to the  operator, blocking transcription  initiation.

·       The trp operon is subject to another layer of regulation.

·        In addition to being controlled at the level of Transcription initiation by the trp repressor, expression of the trp operon is also controlled at the level of transcription elongation by a process called attenuation.

ATTENUATION:

   There are two decision points involved in transcription control

·       The initiation of transcription

·       Continuation of transcription into the operon’s structural genes

·        This additional level of control serves to adjust transcription,such that the two systems of control can decrease transcription more than either one alone. When the repressor protein is not active, RNA polymerase begins transcription of the leader. However, it often does not progress to the first structural gene in the operon. Instead, transcription is terminated within the leader; this is called attenuation.

formation of pause and termination loop - results in no translation

          

·       The trp operon leader contains a sequence of nucleotides called trp L.

·       Interestingly, a small portion of trpL is transcribed and translated, giving rise to a small peptide called the leader peptide.

·       The leader peptide has never been isolated, presumably because it is rapidly degraded. In addition to encoding the leader peptide, trpL contains attenuator sequences.

·       When transcribed, these sequences form stem-loop secondary structures in the newly formed mRNA. We define these sequences numerically (regions 1, 2, 3, and 4).

·       When regions 1 and 2 pair with one another, they form a secondary structure called the pause loop, which causes RNA polymerase to slow down.

·        The pause loop forms just prior to the formation of the terminator loop, which is made when regions 3 and 4 base-pair.

·       A poly-U sequence follows the 3:4 terminator loop, just as it does in factor-independent transcriptional terminators.

·       However, in this case, the terminator is in the leader, rather than at the end of the operon.

·       Another stem-loop structure can be formed in the mRNA by the pairing of regions 2 and 3. The formation of this anti-terminator loop prevents the generation of both the 1:2 pause and 3:4 terminator loops.

·       In the first, translation is not coupled to transcription because protein synthesis is not occurring.

·       In other words, no ribosome is associated with the mRNA. In this, the pause and terminator loops form, stopping transcription before RNA polymerase reaches the trpE gene.

·        In next case translation and transcription are coupled; that is, a ribosome initiates synthesis of the leader peptide.

·       The interaction between RNA polymerase and the  ribosome determines which stem-loop structures are formed. As a ribosome translates the mRNA, it follows the RNA polymerase.

STALLING OF RIBOSOME: (when very less amount of tryptophan is present)

very low level of  trp-2:3 anti-terminatior loop formation
both transcription and translation occurs

            

·       Among the first several nucleotides of region 1 are two tryptophan (trp) codons; this is unusual because normally there is only one tryptophan residue per 100 amino acids in E. coli proteins. If tryptophan levels are low, there will not be enough charged tRNA-trp to fill the A site of the ribosome when the ribosome encounters the two trp codons, and it will stall.

·       Meanwhile RNA polymerase continues to transcribe mRNA, moving away from the stalled ribosome.

TRANCRIPTION OF  Trp GENES:

·        The presence of the ribosome on region 1 prevents region 1- 2 base pairing. As RNA polymerase continues, region 3 is transcribed, enabling the generation of the 2:3 anti-terminator loop. This prevents the formation of the 3:4 terminator loop.

·       The terminator loop is not  formed, RNA polymerase is not ejected from the DNA and transcription continues into the trp structural genes.

FORMATION OF PAUSE AND TERMINATOR LOOP:

·        On the other hand,  when there is plenty of tryptophan in the cell, there will be an abundance of charged tRNA-trp, and the ribosome will translate the two trp codons in the leader peptide sequence without hesitation. Thus the ribosome remains close to the RNA polymerase.

 high level of trp
transcription is terminated while translation of leader peptide occurs

            

·       As RNA polymerase and the ribosome continue through trp L, regions 1 and 2 are transcribed and readily form a pause loop.

·        Then regions 3 and 4 are transcribed, the terminator loop forms, and RNA polymerase is ejected from the DNA template.

·       Finally, the presence of a UGA stop codon between regions 1 and 2 (i.e., the end of the leader peptide-coding region) causes termination of translation too.

also read lac operon -  https://bio-seeks.blogspot.com/2020/06/lac-operon-lac-lactose-operon.html

REFERENCE:

       Prescott's Microbiology by Willey, Sherwood, Wolverton.

     "DON'T BE PUSHED BY YOUR PROBLEMS;

            BE LED BY YOUR DREAMS"