Trp Gene Regulation

Question 1

trp operon expression is controlled at which two points?

Answer 1

-transcription initiation (very similar to lac operon)
-transcription termination

Question 2

mutations in two different sites can lead to constitutive expression of trp operon. what are these two sites?

Answer 2

-trpO (operator site, overlaps promoter)
-trpR (encodes repressor protein that binds to operator & prevents RNAP from transcribing)

Question 3

when does TrpR bind operator? (opposite to LacI)

Answer 3

when tryptophan is present

Question 4

tryptophan is the ____ and trpR is _____ (type of repressor)

Answer 4

-co-repressor
-aporepressor

Question 5

how does tryptophan binding to TrpR affect Trp operon expression?

Answer 5

binding of tryptophan to TrpR changes its conformation so it can bind DNA and repress operon expression

Question 6

In the absence of tryptophan, how does trp operon work?

Answer 6

-trpR gene produces aporepressor protein but no tryptophan means it can’t bind to DNA (trpO) so transcription of trp operon continues (operon on)

Question 7

In the presence of tryptophan, how does trp operon work?

Answer 7

-trpR gene produces aporepressor protein. tryptophan present bind to protein and allows it to bind to DNA (trpO) so transcription of trp operon stops (operon off)

Question 8

catabolic pathways aka _____. anabolic pathways aka _____ (type of operon)

Answer 8

-breaking down/degradative operon
-building things up/biosynthetic operon

Question 9

degradative operon: function? expression? regulation by? effector molecules? presence of effector causes?

Answer 9

-degrade compound
-induced by compound
-repressor
-inducer
-repressor loses DNA binding ability

Question 10

biosynthetic operon: function? expression? regulation by? effector molecules? presence of effector causes?

Answer 10

-synthesize compound
-repressed by compound
-repressor
-corepressor
-repressor gains DNA binding ability

Question 11

repressors of anabolic pathways exist in 2 states. what are they?

Answer 11

aporepressor: repressor protein in absence of effector/corepressor
co-repressor: effector molecule that binds to aporepressor & facilitates DNA binding

Question 12

what does the trp operon encode?

Answer 12

-enzymes for synthesizing l-tryptophan (a.a that is part of most proteins, need to made if not present)

Question 13

trp operon contains what?

Answer 13

5 genes transcribed from single promoter P-trp

Question 14

where the TrpR repressor encoded?

Answer 14

separate place in genome, unlinked (not in operon)

Question 15

trp operon activity when tryptophan is high? low?

Answer 15

-operon not expressed
-operon is turned on

Question 16

contro; of trp operon expression at transcription termination allows for what?

Answer 16

finer control of transcription in proportion to [tryptophan] aka attenuation (controls gene expression @ intermediate levels)

Question 17

how does attenuation work?

Answer 17

transcription start normally but is terminated before RNAP reaches 1st structural gene of operon (happens in operons that have leader region that encodes leader peptide)

Question 18

why does attenuation occur (hint :ribosome)

Answer 18

occurs because ribosome movement during translation affects 2 structure of mRNA

Question 19

3 genetic evidence for 2nd control mechanism/attenuation of Trp operon

Answer 19

• in trpR mutants, higher levels of trp enzymes when tryptophan is not present
  -mutations that lower levels of tRNA-Trp increase trp operon expression
  -deletion of trp leader region eliminates this regulation

Question 20

attenuation of trp operon model (brief)

Answer 20

presence of excess tRNA-Trp, transcription of the trp operon stops in leader region

Question 21

description of trp leader region

Answer 21

-leader peptide with 4 regions of sequence (1,2,3,4) that can form alternative hairpin structures (1:2 & 3:4 or 2:3); short peptide encoded in leader that contains to trp residues

Question 22

How does the trp leader region cause attenuation?

Answer 22

-leader region of mRNA can form intrinstic transcription termination structure; terminator only forms when [try] is high in cell due to different rates of leader peptide translation

Question 23

how to leader peptide translation initiated?

Answer 23

RNAP initiates translation of P-trp. After region1 & 2 of leader are transcribed, stem loop forms, then RNAP pauses and ribosome then initiates leader peptide translation

Question 24

when tryptophan is sufficient, how does attenuation occur after leader peptide translation?

Answer 24

-tRNA-trp is a lot so ribosome translates to end of leader peptide; ribosome at stop codon blocks pairing of segment 2 & 3 of leader mRNA & segment 3 & 4 of leader mRNA form terminator hairpin = attenuation, producing 140 bp transcript (RNA pol terminates)

Question 25

when tryptophan is insufficient, how does attenuation occur after leader peptide translation?

Answer 25

tRNA-trp wil;l be low; ribosome pauses at adjacent trp codons so segment 2 & 3 of leader mRNA can form hairpin (antiterminator) before segment 4 is transcribed ; blocks pairing of 3 & 4 (terminator hairpin). so no transcription termination, and remainder of trp operon transcribed

Question 26

3 pieces of genetic evidence for attenuation model

Answer 26

-mutations that prevent formation of 2:3 hairpin terminate transcription in leader region constitutively
-mutation of AUG start codon of leader peptide yields constitutive transcritipn in leader region (no ribosome stalled on Trp regions, so hairpin1:2 stays, preventing 2:3, so 3:4 forms and stop transcription)
-bacteria starved of other a.a in arg leader peptide also show attenuation