Test Friday 11/16
Chap 16: Regulation of Gene expression in Prokaryotes
- Gene Expression: Transcription - process of converting DNA to RNA
- The cell regulates transcription and can turn on or off the transcription of a gene. It doesn't want to make something it won't use. (sensitive to how they spend energy)
- In prokaryotes genes are typically found by themselves, a single gene OR as operons.
- Operons - a group of genes which are transcribed as a single unit.
- Typically the genes found in an operon are all involved in some process together (this way they can all be regulated together)
- Promoter - upstream of a gene (closer to 5 prime end) site where RNA POL binds to start transcription.
- In prokaryotes there is a sigma factor - a small protein that assists RNA POL in recognizing and binding a promoter.
- After that RNA POL begins copying DNA into RNA and continues until it reaches the Terminator (which is at the end of the gene or operon) and signals the end of transcription. No more than that is copied - don't want to waste energy.
(8:20) How do we regulate the above process of transcription. The study of gene regulation in prokaryotes is EXTENSIVE.
- 1900 it was recognized that cells fail to produce the enzymes for lactose metabolism when lactose is absent. Gave rise to the idea that gene expression is adaptive.
- Constituitive expression - a gene that is always expressed at a relatively high level.
- Positive Regulation - the turning on of the expression of a gene. (Gene is off and then you do something that induces the expression of that gene)
- Negative Regulation - the gene is being expressed until you turn it off. (ex. Tryptophan)
(14:15) Example of Positive Regulation: Lac Operon
- Late 1940's - Jacob + Monod (links to Wikipedia article): laid the groundwork for all the understanding of gene regulation.
- Based on the idea that in a prokaryote the enzymes for lactose metabolism are off without lactose. Their goal was to understand this idea.
- lac operon: three structural genes involved in lactose metabolism
- lac Z - codes for beta galactosidase which breaks down lactose to produce glucose and galactose. You don't want to produce this if you have no lactose.
- lac Y - produces an enzyme called permase which is responsible for facilitating the entry of lactose into the cell.
- lac A - codes for transacetylase. It is believed that it helps to breakdown some of the toxic byproducts of lactose metabolism.
- cis-acting element - a DNA sequence that is bound and acted upon to allow the regulation of a genes expression (turns it on or off).
- trans acting factor - a molecule, often a protein, that binds a cis-acting element to regulate the expression of the gene.
- (24:00) Gratuitous inducer (found by Jacob and Monod) - a molecule which mimics the activity of a molecule which normally activates a system.
- Lactose induces the expression of the lac operon. The inducer can fill the role of lactose.
- IPTG (part of the gratuitous inducer) - when added to the system turned on the expression of the lac operon. This allowed them to find constitutive mutants (mutants which always express the lac operon).
- Lac I - gene upstream of the lac operon (not part of the operon). Lac I is repressor gene that produces a protein which bound and repressed the lac operon. When you mutate the lac I gene, cells consituitively express the lac operon. (Trans acting factor - something that is produced that acts upon a cis-acting element) It no longer represses the expression of the lac operon.
- Lac O - mutation in the operator sequence. Cis acting element. DNA sequence bound by Lac I to prevent transcription of the lac operon. (this is a constituitive mutant)
- Finding the lac I and lac O mutants allowed them to develop a hypothesis as to how the lac operon worked
(33:10) How the regulation of the lac operon works in the absence of sugar.
- The lac I gene produces the Lac I protein. The repressor protein binds to lac O (operator sequence)
- When this happens RNA POL binds the promoter. Repressor protein blocks RNA POL from copying the lac operon, therefore, operon repressed.
How the regulation of the lac operon works in the presence of lactose
- lac I gene produces the Lac I repressor protein. Lactose binds to the Lac I repressor protein. This triggers a confirmation change in Lac I.
- Because of this Lac I cannot bind the operator. SO when this happens RNA POL can bind the promoter and copy the lac operon. The result is that the lac operon is induced.
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