What is the goal of metabolic regulation?
Efficient use of resources
What are the types of metabolic regulation?
-increase/decrease amount of protein
-Increase/decrease activity of protein by preventing functioning of protein
What are the stages of transcription/translation that gene regulation and post-translational modification can take place?
-transcription level w/ regulatory proteins
-transcription or translation level w/ RNA based regulation
How do we study gene expression at the protein level and at the gene level?
Protein level - reporter genes
Gene level - DNA sequencing
What are reporter genes?
Genes that code for proteins that are easy to detect/measure
-eg. green fluorescent protein
DNA sequencing is used to study gene expression at the gene level. How do we do it?
1. Isolate mRNA from cells
2. Transform mRNA to DNA
3. Determine identity and number DNA Sequences
What are regulatory proteins? What kind of structure do they have?
proteins that bind to DNA They have homodimeric structure that bind in major groove in DNA to interact w/ specific DNA sequences
What is the function of a regulatory protein?
Either START or STOP transcription
-block RNA polymerase=prevent transcription
-Bind RNA polymerase=activate transcription
What are the types of transcription regulation?
-repression of mRNA synthesis
-activation of mRNA synthesis
How does negative control of transcription work?
When regulatory protein bound to DNA
-Blocks RNA polymerase and prevents transcription
When regulatory protein released from DNA
-transcription possible but not automatic
How are enzymes used in negative control?
-product abundant in cell=no additional synthesis
-substrate present in cell=additional synthesis
Go through arginine synthesis example, arg operon, for negative control repression.
Arginine acts as co-repressor
-when abundant in cell, it binds repressor and blocks transcription
-when not abundant in cell the repressor doesn't block RNA polymerase
Go through lactose degradation example, lac operon, for negative control induction
lactose acts as inducer
When lactose not abundant in cell the repressor binds DNA to block RNA polymerase
-no need for enzyme
When lactose abundant in cell it binds to repressor to release DNA and allow RNA polymerase to proceed
-need enzyme to break down lactose
What happens if ariginine is high or low in cell? What kind of negative control is this?
What happens if lactose is high or low in cell? What kind of negative control is this?
ARGININE - Repression
LACTOSE - Induction
What is positive control?
When regulatory proteins bound to DNA
-binds RNA polymerase
When regulatory proteins released from DNA
-transcription possible but NOT automatic
Go through maltose catabolism, mal operon, example of activation positve control
Excess maltose in cell = time to break it down
Limited maltose in cell = no need for enzymes
Maltose Activator protein + Inducer
-Maltose activator protein and inducer bind allowing the complex to bind to activator binding site allowing transcription to proceed
What is the difference between postive and negative control?
-control DNA region = activator-binding site
-located BEFORE promoter
-control DNA region = Operator
-Located AFTER promoter
What is the activator-binding site?
Near the promoter region, where activator protein binds to allow transcription to proceed
-if activator binding site distant from promoter then requires DNA to loop
What are activator proteins?
help RNA polymerase recognize the promoter region and begin transcription
What is global control?
single proteins that regulate the expression of multiple operons
-activate/inhibit entire sets of genes
How do E. coli cells choose glucose over other sugars as an energy source?
Single repressor protein, regulons, prevent transcription of group of genes to metabolize other sugars
-alot of glucose present=prevent transcription of other sugar enzymes
-not alot of glucose=allow transcirption of these enzymes
How do microbes sense/respond to environmental conditions?
What are the types of signal molecules that cause microbes to respond to their environment?
-small molecules that permeate membrane
-directly affect transcription
-large molecules that can't permeate membrane
-require signal transduction to affect transcription
What is an example of internal signals?
-gene expression requires minimum number of bacteria to be present
-Release autoinducer signal molecules that activate transcription of genes
How do external signal molecules work?
-external signal activates gene expression
Two component regulatory system
-sensor kinase protein in cytoplasmic membrane detects signal and phosphorylates response regulator
-response regulator binds DNA and represses transcription
How can RNA regulate gene expression?
No regulatory proteins required
What is attenuation?
occurs AFTER initiation of transcription and BEFORE its completion
Forms alternative structures of RNA
-Form 1 = stem+loop form ahead of RNA polymerase
-Form 2 = stem+loop form behind RNA polymerase
What is an example of attenuation?
What are riboswitches?
Form of RNA-based gene regulation that prevents translation
-signal metabolite binds directly to mRNA
-uses secondary mRNA structures like attenuation
What is antisense RNA?
Form of RNA-based regulation
-prevents translation where small RNA, sRNA, binds to synthesized mRNA
-makes double stranded mRNA=no translation b/c cell stops it since only viruses have dsRNA