week 9 (gene exp. in bac.) Flashcards
name: types of gene exp. in bac. (3)
-
constitutive transcription
- constant exp. of genes
⤷ no regulatory control -
regulated transcription
- exp. only occurs under certain conditions -
posttranscriptional regulation
- abundance of mRNA can be modified to influence amount of prot. translated
explain: negative control of transcription
- repressor prot. bind to regulatory DNA seq.
- prevents transcription of a gene
explain: positive control of transcription
- activator prot. binds to regulator DNA seq. and initiates transcription
- direct activation of transcription
name + explain: effect of inducer and corepressors on negative control
-
inducer
- no inducer = repressor blocks transcription
- yes inducer = binds to repressor -> allows release from DNA -
corepressor
- no corepressor = repressor releases from DNA
- yes corepressor = repressor blocks transcription
name + explain: effect of effectors and inhibitors on positive control
-
effector
- no effector = no transcription
- yes effector = binds to activator prot. -> transcription activates -
inhibitor
- no inhibitor = transcription activated
- yes inhibitor = inhibits activator prot. -> no transcription
**inhibitors and effectors bind to effect beha. of activator prot.
explain: negative vs positive control on lac operon
- negative = repressors binds to operator seq -> no transcription
⤷ prevents synth of enz. involved in lactose metabolism
⤷ only activates when lactose is present - positive = transcriptional elements more active when glucose is absent
question: should lactose and glucose be present or absent for the upregulation of the lac operon?
- glucose absence
- lactose present
explain: lactose and e.coli
- glucose = primary E source
- lactose only used when glucose = absent
- lactose needs to be transported into cell by permease transport prot.
- lactose broken down by beta-galactosidase
⤷ into glucose + galactose
⤷ also breaks into allolactose
explain: role of allolactose
- induced compound
- binds to repressor prot. -> removes inhibition of lac operon -> allows transcription
name: order of struc. in lac operon (which parts of repressor vs promoter vs operator)
CAP binding site -> lacP -> lacO -> lacZ
CAMP, lacP, lacO = promotor region
**P = promotor seq.
O = operator
**lacI is before the lac operon + acts as regulatory gene (not considered part of the operon)
question: where on the lac operon are beta-galactosidase, permease, and transacetylase?
beta = lazZ
permease = lacY
⤷ allows more lactose of enter
transacetylase = lacA
⤷ thought to protect cell against harmful enz.
explain: the CAP-cAMP complex
- positive control of lac operon
- CAP upregulates transcription by RNA poly
- cAMP = synthesized by adenylate cyclase
⤷ glucose inhibits adenylate cyclase - low glucose -> high cAMP -> forms complex -> transcription of lac operon
- high glucose -> low cAMP -> no complex -> no transcription
question: what happens to lac operon in presence of glucose and absence of lactose? vv? both present?
YES GLUC NO LAC
- no activation of lac operon transcription
- cAMP low -> no CAP-cAMP complex -> no upregulation
- also no lactose = no allolactose -> repressor stays on
NO GLUC YES LAC
- repressor removed by allolactose
- RNA poly binds to promotor -> transcription
- cAMP high -> recruits more RNA poly.
BOTH
- cAMP still low
- low transcription
- repressor still removed
⤷ bc still allolactose
⤷ basal lvl
name: lac operon mutations (3)
loss of function in lacI
- constitutive expression of lac operon (always on)
operator constitutive mutation
- prevents WT repressor from binding
- constitutive expression of lac operon
super repressor mutation
- prevents inducer from suppressing repressor (repressor always bound to operon)
- no transcription of lac operon
question: how to check if a plasmid was inserted correctly?
- plasmid into cell
- grow cell on plate w/ antibiotics
- only the cells w/ plasmid properly in can have antibiotic resistance
⤷ means only the cells that can grow had it inserted
question: how to control the expression of a gene using the lac operon?
- put gene of interest downstream of lac operon
- insert plasmid
- repressors can prevent expression of inserted gene
explain: trp operon structure
- 5 genes (order):
⤷ E, D, C, B, A
⤷ each gene catalyzes a step in synthesizing tryptophan - promoter trpP, operator trpO, leader trpL all upstream
- repressor = outside operon
⤷ trpR
question: where would a repressor bind on the trp operon to inhibit the promotor?
- trpO
explain: regulation of trp operon by tryptophan
- higher trp conc. -> more binds to repressors -> more inhibition
- trp absent -> repressor no longer binds -> allows synthesis of trp
**trp present still allows some transcription but low (basal)
explain: regulation of trp operon by attentuation
- controlled by hairpin structures from trpL mRNA
⤷ trpL = leader (upstream) - dep. on lvls of trp
- hairpin between regions 3 and 4 -> no transcription
⤷ caused by rapid translation of trpL bc lots of trp available - hairpin between regions 2 and 3 -> transcription
⤷ caused by slow transcription of trpL bc low trp lvls
explain: heat shock
- transcriptional activation triggered by heat
- in e.coli = higher than 37 degrees
explain: heat shock response in e.coli
- activated by higher than 37 degrees
⤷ use alternative sigma factors to activate heat shock response gene - uses chaperone prot. to help fold or degrade prot. damaged by heat
