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Prokaryotic Gene Regulation: Lecture 5. Introduction The two types of transcription regulation control in prokaryotic cells The lac operon an inducible.

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Presentation on theme: "Prokaryotic Gene Regulation: Lecture 5. Introduction The two types of transcription regulation control in prokaryotic cells The lac operon an inducible."— Presentation transcript:

1 Prokaryotic Gene Regulation: Lecture 5

2 Introduction The two types of transcription regulation control in prokaryotic cells The lac operon an inducible regulatory feedback system The tryp operon a repressible regulatory feedback system. The role of bioinformatics analysis in such systems

3 Gene Regulation All genes must have some way of regulating their expression (converting the DNA to amino acids) in order to allow them to adopt appropriately to the environment. In prokaryotic cells the process, owing to the simple nature of the genomic material, is basically controlled mainly at the transcription level… Essentially the molecule RNA polymerase must bind to an exposed part of DNA strand called a promoter; it must then move, in the 5 to 3 direction, transcribing the DNA sequence of the gene to RNA. – The transcribed sequence begins at the Transcription start site (TSS) and finishes at the Transcription Termination site (TES) ; – The sequence of DNA that is translated into the amino acid sequences is knows as the CDS (coding sequence)

4 Types of Transcription control The transcription is controlled/ regulated at two stages: – RNA polymerase binds to the DNA strand; If it binds to DNA transcription begins Else if it is prevented from binding; DNA is not transcribed: gene not expressed. – RNA polymerase moves in the 5 to 3 direction: If the movement of RNA polymerase is [physically] blocked transcription is stopped and gene is not expressed Otherwise transcription is completed and the gene is expressed.

5 Transcriptional control systems Inducible: gene is expressed (DNA is converted to RNA) only if the molecule (inducer) is present. Repressible: if molecule is present gene expression is turned off In both cases the molecule works via a feedback loop.

6 gene expression regulatory loops Feedback loop: product of the gene expression loops back (directly/indirectly) and alter the expression of the same gene. The effect can be either : – positive : turns on transcription via an inducer; e.g. [lactose ] – Negative: turns off transcription via a repressor e.g. [tryptophan] Gene Gene product

7 Inducible Prokaryotic regulation Lactose, a complex sugar (glucose) In order for E. Coli to use (metabolise) the sugar a gene system referred to as the lac operon must produce three enzyme(s) that allow lactose to be utilised by the bacteria. For simplicity we will refer to the combined system as: lactose dehydrogenise (a more detailed description of the enzymes and their functions can be found in Klug p. 310). The function of this enzyme is to break convert lactose to glucose and increase its absorption into the bacterial cell. Basically In order to ensure efficient functionality of the lac operon system it must have the following properties: – will not be expressed if there is no lactose – will be expressed if there is lactose. Note a more complete description of the system, involving glucose, can be found in the supplementary notes.

8 Function of Lac operon The term operon is the common gene regulatory system used in prokaryotic cells and generally a number of genes are regulated as a one. In the E Coli Lac operon DNA there are: – 1 repressor gene ( – A promoter (where RNA polymerase binds) – A Cis–acting regulatory region (operator) – A leader region (where transcription starts but leader is an UTR) – 3 structural genes: LacZ, LacY and LacA (refer to here as lactose dehydrogenase); actually 3 different genes with different functionality in the utilisation of lactose. Klug chapter 15

9 Function of Lac operon RNA polymerase binds to the promoter region The repressor gene produces a product a repressor protein This binds to the DNA at the operator region and blocks RNA polymerase moving down the DNA strand. If lactose is present it alters the repressor protein. The alter repressor protein is unable to bind to the DNA RNA polymerase binds to the promoter region and begins transcribing the 3 structural genes. When lactose levels drop to zero: what happens? Klug chapter 15 RNA polymerase Repressor protein

10 Glucose and the lac operon Lactose is metabolised into glucose so what happens if glucose is present. Catabolite-activation protein (CAP): CAP must be present to make RNA polymerase binding efficiently In the presence of glucose the CAP is altered and prevents RNA polymerase binding to the promoter region and so prevents transcription. Klug chapter 15

11 A repressible operon Tryptophan is an essential AA and is normally made (biosynthesised) by E Coli. If tryptophan is present externally [in sufficient levels] then the biosynthesis stops; The DNA strand has the same elements as the lactose operon: repressor gene, promoter, operator, leader and genes In this system the repressor protein is altered by tryptophan and the modified repressor protein now binds to the operator region and blocks RNA polymerase transcribing the genes required to make tryptophan. Klug chapter 15

12 The tryptophan operon In addition in the presence of tryptophan there is an additional control mechanism called: The attenuation regulatory mechanism: In the sequence prior to structural genes is the attenuator region: If tryptophan and its gene expression is repressed they still found that transcription was initiated… ; there was RNA fragments of leader [L]sequence: this shows that transcription begins at a transcription start site (TSS) upstream of AUG (start codon) Thus altering the repressor protein is not enough to prevent expression. It seems that tryptophan also binds to the attenuator [A]region and prevents transcription beyond the leader region. Attenuator region Leader region Klug chapter 15

13 Importance of bioinformatics Bioinformatics can help with improving our understanding of such regulation by: – Finding potential gene regions and promoter regions since a gene will be in close proximity to a promoter regions. – The prokaryotic sequence normally has specific sequences associated with it and so do genes [begin with AUG/ATG]. [This will be covered in more detail in the lecture finding genes/promoters – Prokaryotic systems usually have polycistronic mRNA ; multiple gene sequences in very close proximity (contiguous) or in some cases the may even slightly overlap. (understanding bioinformatics section 9.2) – The gene sequence can be converted into amino acid sequence via universal code – Finally the promoter sequences and gene sequences can be analysed using advanced computational techniques to help understand or suggest how the operon works

14 Exam question Distinguish, using suitable examples, the difference between inducible and repressible transcription control in prokaryotic organisms. (20 marks) Discuss, using examples, how analysis of DNA sequences can help improve our understanding of genome in prokaryotic cells. (10 marks)

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