1.8 - Bacterial Transcription + Lac Operon Flashcards
sigma (σ) factor
polymerase subunit that recognises and binds to promoter sequences on DNA
result of sigma factor and RNA polymerase combining
holoenzyme
what does binding of sigma allow RNA polymerase to do?
recognise transcription site and strongly bind to DNA
what do multiple sigma facors allow for?
different groups of genes to be regulated
most abundant sigma factor
sigma 70 (σ70)
role of sigma 70 (σ70)
responsible for transcription of housekeeping genes
regulator sites
located around promoter, can inhibit or enhance transcription
operator site
sequence recognised by repressor genes
effect of repressor genes binding to operator site
physically blocks RNA polymerase from binding
activator sequences
bind proteins that can help to increase efficiency of RNA polymerase to bind to DNA
“melting” of DNA helix
sigma factor within holoenzyme initiates “melting” of DNA helix (separation of strands to prepare for transcription)
what determines whether top/bottom DNA strand is transcribed?
which strand the specific promoter sequence that sigma factor has bound to is on
when can transcription of the RNA begin?
when DNA is melted and strands exposed
what replaces thymin in RNA transcript?
uracil
when does sigma factor disassociate?
as RNA transcript is being created (no longer needed) - free to bind to other RNA polymerase to initiate transcription elsewhere
elongation
RNA polymerase continues to add nucleotides to end of RNA transcript until signalled to stop transcribing at termination sequence
terminator sequence (2)
- complementary sequences of this creates hairpin structure, causes RNA polymerase to stall
- combined with stretch of uracils on end of transcript, there is reduced interaction between DNA, RNA and RNA polymerase
(causes termination of transcription and release of RNA transcript)
effect of transcription occurring in cytoplasm (bacteria)
most of the time, new transcripts exposed to proteins in cytoplasm, can include ribosomes which carry out translation
result of bacterial transcripts being exposed to ribosomes in cytoplasm
transcription and translation can occur simultaneously, means RNA transcript generally not modified
why do bacteria require tight transcriptional control?
single cell organisms, ensures efficient use of energy and resources
operons
polycistronic genes allowing for coordinated expression of multiple genes at same time
what are operons under control of?
the same promoter
Lac operon
multiple genes that form Lac Operon all involved in metabolism of lactose into glucose and galactose
genes within Lac Operon (3)
- lacZ gene - β-galactosidase
- lacY gene -galactoside permease
- lacA gene - β-galactosidase transacetylase
default state of Lac Operon
“turned off” through Lac repressor coded by lacI gene (lower case lac, upper case I), occurs when cell has the glucose it needs
how is the lacI gene a trans-regulator of the Lac Operon?
has its own promoter and terminator sequence
state of Lac repressor (lacI)
always transcribed and expressed
role of lac repressor
bonds to operator in Lac Operon preventing RNA polymerase from binding
allosteric regulation
regulatory mechanism whereby proteins interacting with sequences often require additional molecules to allow them to bind to DNA
how do additional molecules allow DNA-binding proteins to bind (allostery)
bind to DNA-binding protein causing conformational change altering how it binds to DNA
what does allostery allow for?
activation or inhibition of gene expression in response to stimuli
different effects of allosteric regulation (dependant on DNA-binding proteins) (4)
- inducing repressors
- repressible repressors
- repressible activators
- inducible activators
inducing repressors (allostery)
do not inhibit transcription when bound to effector molecule
repressible repressors (allostery)
inhibit transcription when bound to effector molecule
repressible activators (allostery)
do not activate transcription when bound to effector molecule
inducible activators (allostery)
causes transcription when bound to effector molecule
binding of CAP to Lac Operon (3)
- when cell runs out of glucose, cell produces cyclic AMP from breaking down ATP
- cyclic AMP can bind to protein cAMP-bound catabolite activator protein (CAP)
- cyclic AMP acts as inducible activator of CAP, can then bind to CAP binding site upstream of Lac Operon promoter
activity of CAP if there’s no lactose for the cell to convert into glucose
repressor remains bound to DNA to prevent CAP from helping to bind RNA polymerase
(no point wasting further energy making enzymes when no lactose to break down)
role of allolactose (2)
- if lactose present in cell it will be metabolised into allolactose by β-galactosidase (always small amount in cell)
- allolactose acts as inducible repressor, binds to Lac repressor protein, causing conformation change and can no longer bind to operator site
(detector system for presence of lactose)
result of removal of Lac repressor
RNA polymerase with sigma factor able to bind to DNA, transcription and subsequently translation occur producing protein, this allows lactose to be transported into cell and broken down into glucose
activity of CAP when returning to baseline (glucose present) (4)
- cyclic AMP reduces as glucose becomes main energy/carbon source, meaning less cAMP to bind to CAP
- means less CAP able to bind to DNA
- ensures cell not continuously making more enzyme where glucose/lactose present (already converted as needed)
- eventually lactose/allolactose levels drop and cycle restarts with binding of Lac repressor to operator
summarise how Lac Operon regulates expression of genes involved in metabolising lactose to glucose
by using repressor and activator, ensures process is down to efficient manner when required without wasting energy making gene products that won’t be used
tryptophan operon
operon where there are five genes within it relating to tryptophan biosynthesis
what regulates binding of active repressor to operator site within promoter region in tryptophan operon
tryptophan levels
active repressor action when there are high levels of tryptophan
acts as repressible repressor and turns off operon
maltose operon action at high maltose levels
maltose binds to maltose activator protein, allowing for expression of genes for maltose metabolism
maltose operon action at low maltose levels
less to bind to activator protein, transcription and gene expression of operon is reduced
