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GENETICS > Lecture 6 cards > Flashcards

Lecture 6 cards Flashcards

Prokaryotic regulation

1
Q

What are the 2 main types of regulatory proteins?

A
  1. activators; 2. repressors
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2
Q

What are the 2 categories of small effector molecules?

A
  1. inducers; 2. inhibitors
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3
Q

What is polycistronic mRNA?

A

mRNA that contains two or more genes

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4
Q

What are the 4 structural components of operons?

A
  1. promoter;2. operator;3.structural genes;4.terminator
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5
Q

What comprises the lac operon?

A

3 regulatory elements:1. promoter (lacP)2. operator(lacO) 3. CAP site;3 structural components: 1. lacZ 2. lacY 3. lacA

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6
Q

What does lacZ encode for and what does it do?

A

B-galactosidase: (1)cleaves lactose and lactose analogues (ONPG) (2) converts lactose to allolactose

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7
Q

What does lacY encode for and what does it do?

A

lactose permease: the membrane component required for lactose transport

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8
Q

What does lacA encode for and what does it do?

A

galactoside transacetylase: modifies lactose and its analogues covalently

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9
Q

What is lacO?

A

lacO is an operator sequence (sometimes referred to as a negative control element) that contains the binding site for the lac repressor (lacI)

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10
Q

What denotes a mutation on the lacO that abrobates binding of lacI?

A

lacO^(c) /////lacO-

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11
Q

What is lacP?

A

lacP is the promoter sequence that contains the recognition/binding site for RNA polymerase?

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12
Q

What denotes a mutation in the promoter sequence of the lac operon that abrogates binding of RNA polymerase?

A

lacP-

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13
Q

What type of mechanism controls the lac operon and what are the 2 regulatory components?

A

the lac operon is most heavily influenced by an inducible negative control mechanism(1)LacI– the repressor protein (2)Allolactose–an inducer

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14
Q

Under what condition is the LacI repressor bound to the operon?

A

no allolactose

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15
Q

What is needed for LacI to be inactivated?

A

allolactose must be bound to LacI

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16
Q

What is the function of allolactase in the lac regulatory system?

A

It binds to the lac repressor and inactivates it via conformational change that prevents LacI from being able to bind to DNA

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17
Q

What is lacI?

A

-lacI is a gene that encodes for the Lac repressor protein;-its promoter is called the i promoter; -constitutive expression at low levels

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18
Q

What are 3 different forms of lacI/LacI?

A

(1) lacI+(2)lacI-(3)lacI^(s)

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19
Q

lacI+

A

Normal regulation: lac operon ON in presence of lactose and OFF in the absence.

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20
Q

lacI-

A

No repressor made: constitutive expression even in absence of lactose –>”recessive”

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21
Q

lac^(s)

A

super-repressor made: lac operon “always” OFF–>”dominant”

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22
Q

Describe the real lacO structure:

A

O1: next to the promoter (downstream);O2: Downstream in the lacZ coding region;O3: slightly upstream of the CAP site

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23
Q

LacI has to bind to ___ of the ___ operators to repress _____.

A

2, 3, transcription

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24
Q

What are the only two combinations of portions of the lacO structure that lacI can bind to?

A

O1 and O2/O1 and O3

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25
Q

How does the LacI bind to the two operator sites?

A

It has to form a loop in the DNA

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26
Q

What does the loop formed by the binding of the LacI repressor protein do?

A

The loop hides the lac promoter and RNA pol cannot bind

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27
Q

What is catabolite repression?

A

a means by which the lac operon can be regulated if lactose and glucose are available.

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28
Q

If lactose and glucose are available, how does catabolic repression accomplish regulation of the lac operon?

A

catabolic repression prevents the use of lactose while glucose is first metabolized. Then, when glucose is depleted, catabolic repression is alleviated and the lac operon is expressed.

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29
Q

What purpose does cyclic AMP (cAMP) serve in catabolic repression?

A

cAMP serves as the effector molecule by binding to the CRP/CAP (cyclic AMP receptor protein/catabolic activator protein)

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30
Q

adenylyl cyclase (cya)

A

enzyme that uses ATP to make cAMP

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31
Q

What complex exhibits positive control and is inducible?

A

the cAMP-CRP complex

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32
Q

What is the function of the cAMP-CRP complex?

A

it increases transcription by interaction with the C-terminal domain of RNA pol alpha subunit aiding in RNAP recruitment/binding to promoter

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33
Q

Where does the cAMP-CRP complex bind?

A

it binds to the CAP site near the lac promoter

34
Q

Describe the affect that glucose has in the role of regulation of the lac operon:

A

When glucose is present adenylyl cyclase (cya) is inhibited which results in the decrease of cAMP which is no longer able to be used to increase the rate of transcription. This ultimately

35
Q

Lactose What does this scenario result in? Why is it ultimately favorable?

A

low rate of transcription because very little cAMP binding. This is a favorable state because the operon is primed for either one of the sugars… it could go either way.

36
Q

What comprises the ara operon?

A

-3 structural genes (BAD):1. araB 2. araA 3. araD; - promoter = P_(BAD); -CAP site

37
Q

What is the role of the CAP site?

A

serves as a binding site for CRP

38
Q

CRP

A

catabolite activator protein

39
Q

araC

A

gene; -encodes the regulatory protein AraC. ; -promoter is called Pc; -this gene is next to ara operon but it is not a part of it!

40
Q

What 3 sites does the AraC protein bind to?

A
  1. araI; 2. araO1; 3. araO2
41
Q

AraC

A

protein; -binds to operator sites of the ara operon; -can act as a “+” or “-“ regulator based on arabinose presence

42
Q

What happen to the ara operon when glucose levels are low?

A

CAP-cAMP activation occurs

43
Q

Describe the ara operon without arabinose?

A

closed latch;–>Tight bend prevents RNA pol from transcribing the ara operon from the P_(BAD) promoter. “an AraC protein binds to the araO2 operator site and another to the araI site. These two AraC proteins interact to promote a loop in the DNA.”

44
Q

How does AraC accomplish the repression of the ara operon?

A
  1. In the abscence of arabinose, AraC binds to araO2 and araI.; 2. The multiple AraC proteins can then interact with each other which then causes the DNA to loop; 3. The loop then blocks RNA pol from accessing the araC promoter [Pbad]
45
Q

Describe the ara operon with arabinose?

A

opened latch; The arabinose binds to AraC proteins which leads to a conformational change that breaks the interaction b/w the AraC proteins bound @ the araO2 & araI operators. This in turn, causes the DNA loop to break. This allows for an AraC dimer to bind to araI which activates transcription!

46
Q

How does AraC accomplish the activation of the ara operon?

A
  1. When arabinose is present it binds to the AraC proteins; 2. This ultimately causes the DNA loop to open.; 3. An AraC dimer then binds to araI and activates transcription!
47
Q

What 2 genes are involved in the regulation of the trp operon?

A

trpL & trpR

48
Q

trpR

A

-encodes for the trp repressor protein. ; -inhibited by tryptophan

49
Q

trpL

A

-encodes a short peptide called the Leader peptide ‘ -The way that the trpL mRNA is translated determines the conditions that dictate the formation of the 3-4 stem loop.

50
Q

Leader Peptide

A

-Encoded by the trpL gene.; -Involved in attenuation.

51
Q

2 ways the trp operon is regulated:

A
  1. repression ; 2. attenuation
52
Q

When can repression of the trp operon occur?

A

before mRNA is made

53
Q

When can attenuation of the trp operon occur?

A

after mRNA has begun to be made

54
Q

Before mRNA is made: What happens to the trp operon when tryptophan levels are low?

A

transcription of the entire trp operon occurs b/c TrpR is inactive

55
Q

Before mRNA is made: What happens to the trp operon when tryptophan levels are high?

A

repression of the trp operon occurs b/c TrpR is active when bound by tryptophan.

56
Q

After mRNA has begun to be made: what happens to the trp operon when there are high levels of tryptophan?

A

attenuation occurs b/c the “sensor” detects low trp levels

57
Q

Where is the Trp “sensor” located?

A

within region 1

58
Q

What occurs in the process of attenuation?

A

when Attenuation occurs, the RNA is transcribed only to the attenuator sequence, and then transcription is terminated.

59
Q

Why is attenuation possible in prokaryotes?

A

this form of regulation is posible in prokaryotes b/c transcription&translation are coupled

60
Q

What does the attenuation of the trp operon accomplish?

A

it inhibits the further production of tryptophan

61
Q

What are the different regions of the trp operon?

A

Regions 1,2,3,4

62
Q

Which regions within the trp operon are complementary to each other?

A

Region 2 : 1 and 3; Region 3 : 2 and 4

63
Q

How is transcription of the trp operon terminated?

A

the formation of the 3-4 stem loop causes RNA pol to pause. Following this loop is the U-rich attenuator which allows for the dissociation of the RNA pol from the DNA.

64
Q

What are the 3 possible scenarios of the translation of the trpL mRNA?

A
  1. no translation; 2. low levels of translation; 3. high levels of translation
65
Q

Scenario 1: What happens if translation is not coupled with the transcription of the trp operon?

A

-The stable form of mRNA includes the H-bonding of regions 1&2 and regions 3&4.; - This results in the termination of transcription just past the trpL gene b/c of the terminator stem-loop that is formed.

66
Q

Scenario 2: How does low levels of trpL mRNA translation occur?

A

[trp] levels are very low: 1. there isnt enough tRNA^(trp) to fill the pair of trp codons in the “sensor” ; 2. the ribosome pauses @ region 1; 3. regions 2 & 3 then form a stem loop; 4. transcription still continues b/c the terminator and the 3-4 stem-loop are not formed.

67
Q

Scenario 3: How does high levels of trpL mRNA translation occur?

A

[trp] levels are high:; 1. enough tRNA^(trp) so translation continues through the sensor until the translation stop codon.; 2. the ribosome pauses @ region 2 ; 3. regions 3 & 4 then form a stem loop; 4. transcription is terminated b/c the terminator and the 3-4 stem-loop are formed

68
Q

What are the 2 ways to regulate translation?

A
  1. translational repressors; 2. antisense RNA
69
Q

What are 2 ways that translational repressors can inhibit translation?

A

1- binding next to the Shine-Dalgarno sequence and/or the start codon; 2- binding outside the Shine-Dalgarno / start codon region

70
Q

How does a translational repressor binding next to the Shine-Dalgarno sequence and/or the start codon inhibit translation?

A

it sterically hinders the ribosome from initiating translation

71
Q

How does a translational repressor binding outside the Shine-Dalgarno / start codon region inhibit translation?

A

this stabilizes an mRNA secondary structure that prevents initiation

72
Q

What is antisense RNA?

A

RNA strand that is complementary to mRNA

73
Q

What are the 2 types of antisense RNA?

A

siRNA and miRNA

74
Q

siRNA

A

antisense RNA; longer; generally perfect matches for a single target

75
Q

miRNA

A

antisense RNA; shorter; a single one can affect multiple RNAs

76
Q

OmpF

A

protein in E.coli; important in osmoregulation; preferentially produced at low osmolarity.

77
Q

How is OmpF at high osmolarity?

A

synthesis is decreased b/c of expression of micF

78
Q

micF

A

doesnt code for a protein BUT it is complementary to ompF mRNA (antisense RNA)

79
Q

What are the 2 posttranslational regulation mechanisms?

A
  1. feedback inhibition; 2. covalent modification of the structure of proteins
80
Q

feedback inhibition:

A

The final product in a pathway can inhibit an enzyme that acts early in the pathway.; Enzyme 1= allosteric enzyme (has 2 different binding sites)

81
Q

irreversible covalent modifications in posttranslational regulation:

A

1-proteolytic processing; 2- attachment of prosthetic groups, sugars, or lipids

82
Q

reversible covalent modifications:

A

-phosphorylation(-PO4); -acetylation(-COCH3); -methylation(-CH3)

GENETICS (3 decks)

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