hebrews 1:1-2
DESCRIPTION
Hebrews 1:1-2 1 God, who at sundry times and in divers manners spake in time past unto the fathers by the prophets, 2 Hath in these last days spoken unto us by his Son, whom he hath appointed heir of all things, by whom also he made the worlds;. Transcription From DNA To RNA. - PowerPoint PPT PresentationTRANSCRIPT
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©2000 Timothy G. Standish
Hebrews 1:1-2 1 God, who at sundry times and in
divers manners spake in time past unto the fathers by the prophets,
2 Hath in these last days spoken unto us by his Son, whom he hath appointed heir of all things, by whom also he made the worlds;
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©2000 Timothy G. Standish
TranscriptionTranscriptionFrom DNA To RNAFrom DNA To RNA
Timothy G. Standish, Ph. D.
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DNA
mRNA
Transcription
IntroductionIntroduction
The Central Dogma The Central Dogma of Molecular Biologyof Molecular Biology
Cell
Polypeptide(protein)
TranslationRibosome
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©2000 Timothy G. Standish
RNA PolymeraseRNA PolymeraseRNA Polymerase is a spectacular enzyme,
functioning in:
1Recognition of the promoter region
2Melting of DNA (Helicase + Topisomerase)
3RNA Priming (Primase)
4RNA Polymerization
5Recognition of terminator sequence
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©2000 Timothy G. Standish
5’ 3’
RNA Transcript
A “Simple” GeneA “Simple” Gene
Protein Coding Region
Terminator Sequence
Promoter/Control Region
3’ Untranslated RegionTranscription Start Site 5’ Untranslated Region
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©2000 Timothy G. Standish
Stages of TranscriptionStages of Transcription
1Initiation
2Elongation
3Termination
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©2000 Timothy G. Standish
Prokaryotic Transcription InitiationProkaryotic Transcription Initiation The subunit of prokaryotic RNA
polymerase is necessary for promoter recognition and binding of RNA polymerase to the promotor
Different subunits allow recognition of different types of promoters thus the type of genes transcribed can be modulated by altering the types of a subunits which attach to RNA polymerase
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©2000 Timothy G. Standish
Prokaryotic Transcription InitiationProkaryotic Transcription Initiation
Constitutive GeneHeat Shock GeneP1 P2
Different promoters
RNAPol.
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©2000 Timothy G. Standish
Prokaryotic Transcription InitiationProkaryotic Transcription Initiation
Constitutive GeneHeat Shock Gene P2P1
Different promoters
RNAPol.
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©2000 Timothy G. Standish
Eukaryotic Transcription InitiationEukaryotic Transcription InitiationProteins called transcription factors bind to
the promoter region of a gene If the appropriate transcription factors are
present, RNA polymerase binds to form an initiation complex
RNA polymerase melts the DNA at the transcription start site
Polymerization of RNA begins
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©2000 Timothy G. Standish
RNA Pol.
InitiationInitiation
Promoter
RNA Pol.T. F.
T. F.
T. F.
5’RNA
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©2000 Timothy G. Standish
RNA Pol.
InitiationInitiation
RNA Pol.
T. F.
5’RNA
Promoter
T. F.
T. F.
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Transcription TerminationTranscription TerminationThere are two types of termination: Rho dependent requires a protein called Rho, that
binds to and slides along the RNA transcript. The terminator sequence slows down the elongation complex, Rho catches up and knocks it off the DNA
Rho independent termination depends on both slowing down the elongation complex, and an AT-rich region that destabilizes the elongation complex
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©2000 Timothy G. Standish
RNAPol.
5’RNA
RNA Pol.
5’RNA
TerminationTerminationRho IndependentRho Independent
Terminator
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©2000 Timothy G. Standish
RNAPol.
5’RNA RNA
Pol.
5’RNA
TerminationTerminationRho IndependentRho Independent
Terminator
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©2000 Timothy G. Standish
RNAPol.
5’RNA
TerminationTerminationRho DependentRho Dependent
Terminator
RNA Pol.
5’RNA The terminator
sequence slows RNA polymerase
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RNAPol.
5’RNA
TerminationTerminationRho DependentRho Dependent
Terminator
Help, Rhohit me!
RNA Pol.
5’RNA
Rho catches up with RNA polymerase
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RNAPol.
5’RNA
TerminationTerminationRho DependentRho Dependent
Terminator
RNA Pol.
5’RNA
The elongation complex disintegrates
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DifferencesDifferencesBetween Transcription InBetween Transcription In
Prokaryotes and Prokaryotes and EukaryotesEukaryotes
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3’
5’
5’
3’
Transcription And Translation Transcription And Translation In ProkaryotesIn Prokaryotes
Ribosome
Ribosome5’
mRNA
RNAPol.
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DNA
Cytoplasm
Nucleus
Eukaryotic TranscriptionEukaryotic Transcription
ExportG AAAAAA
RNA
Transcription
Nuclear pores
G AAAAAA
RNAProcessing
mRNA
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A “Simple” Eukaryotic GeneA “Simple” Eukaryotic Gene
Terminator Sequence
Promoter/Control Region
Transcription Start Site
RNA Transcript
5’ Untranslated Region 3’ Untranslated Region
Exons
Introns
3’5’ Exon 2 Exon 3Int. 2Exon 1 Int. 1
3’5’ Exon 2 Exon 3Exon 1 Int. 2Int. 1
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3’5’ Exon 2 Exon 3Int. 2Exon 1 Int. 1
Processing Eukaryotic mRNAProcessing Eukaryotic mRNA
Protein Coding Region
3’ Untranslated Region5’ Untranslated Region
3’AAAAA
3’ Poly A Tail
5’ G
5’ Cap
Exon 2 Exon 3Exon 1
Int. 2
Int. 1
RNA processing achieves three things: Removal of introns Addition of a 5’ cap Addition of a 3’ tail
This signals the mRNA is ready to move out of the nucleus and may control its lifespan in the cytoplasm
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Common Splicing MechanismCommon Splicing Mechanism
Exon 2Exon 1Intron
AGAGU 3’5’18-40 BP
Branch site
Left (donor)5’ splice
site
Right (acceptor)3’ splice
site
Py80NPy80Py87Pu75APy95 (Animal-Subscripts indicate percent frequency)
U A C U A A C (Yeast)
The branch sequence allows identification of the 3’ splice site
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©2000 Timothy G. Standish
Common Splicing MechanismCommon Splicing MechanismLariat Lariat FormationFormation
Exon 2A AG
GU
3’
5’
3’
Exon 1Intron
Lariat
Yee ha!Lariat
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Common Splicing MechanismCommon Splicing Mechanism
3’
Exon 2Exon 15’
A
AG
GU
Intron lariat
Following excision, the lariat is rapidly degraded
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Common Splicing MechanismCommon Splicing Mechanism
3’
Exon 2Exon 15’
Following excision, the lariat is rapidly degraded
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©2000 Timothy G. Standish
The SpliceosomeThe Spliceosome Spliceosomes are structures that form within the
nucleus to remove introns from eukaryotic hnRNA This structure is large, on the order of a ribosome
subunit Like the ribosome, spliceosomes are composed of
both protein and RNA
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Wobble Base Wobble Base PairingPairing
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©2000 Timothy G. Standish
The Rules of Codon The Rules of Codon Anticodon Base PairingAnticodon Base Pairing
Three things affect the way in which base pairing occurs between codons on mRNA and anticodons on tRNA:
1 How the two molecules “twist” when annealing - They are not free to form a perfect A helix
2 The environment of the Ribosome A site
3 Chemical modification of bases These three factors change the usual base pairing
seen in DNA and RNA, particularly at the first base of anticodons/third base of codons
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U*
9
262223Pu
16
12Py 10
25
20:1
G*
17:1
Pu
A20:2
1713
20G
A5051
656463
G
62
52
CPu
59
A*
C
Py
T49
39
4142
31
2928
Pu*
43127
U35
38
36
Py*
34
403047:1
47:15
46
Py47:16
4544
47
73CCA
7071C
66676869
32G
7654
D Arm - Contains dihydrouridine
Acceptor Arm - A specific amino acid is attached to the 3’ end
TC arm - stands for pseudouridine
Extra Arm - May vary in size
Transfer RNA (tRNA)Transfer RNA (tRNA)3’
5’
Anticodon
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©2000 Timothy G. Standish
U*
9
262223Pu
16
12Py 10
25
20:1
G*
17:1
Pu
A20:2
1713
20G
A5051
656463
G
62
52
CPu
59
A*
C
Py
T49
39
4142
31
2928
Pu*
43127
U35
38
36
Py*
34
403047:1
47:15
46
Py47:16
4544
47
73CCA
7071C
66676869
32G
7654
D Arm - Contains dihydrouridine
TC arm - stands for pseudouridine
Transfer RNA (tRNA)Transfer RNA (tRNA)
Pseudo-uridine
N ON
O
HN NH
Dihydro-uridine
O
O
NHHH
HH N
Anticodon
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©2000 Timothy G. Standish
Some Other Strange tRNA BasesSome Other Strange tRNA Bases
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NO
H
NO
N
NH C
ytosine
H
O
NN
N
N
N
H
H
Guanine -+
+
+
-
-
Base PairingBase PairingGuanine And CytosineGuanine And Cytosine
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N
O
N
ON
H+
- UracilN
NN
N
HN H
-
+Adenine
Base PairingBase PairingAdenine And UracilAdenine And Uracil
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Base PairingBase PairingAdenine And CytosineAdenine And Cytosine
NO
H
NO
N
NH C
ytosine-
+
-
N
NN
N
HN
H
-
+
Adenine
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N
O
N
ON
H+
- Uracil
Base PairingBase PairingGuanine And UracilGuanine And Uracil
H
O
NN
N
N
N
H
H
Guanine
+
+
-
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NO N
O
NHUracil
+
-
Wobble Base PairingWobble Base PairingGuanine And UracilGuanine And Uracil
H
O
NN
N
N
N
H
H
Guanine
+
+
-
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N
S
N
ON
H+
-2 Thio-
uracilN
NN
N
HN H
-
+Adenine
Base PairingBase PairingAdenine And 2-ThiouracilAdenine And 2-Thiouracil
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S N
O
NH
Wobble Base PairingWobble Base PairingGuanine And 2-ThiouracilGuanine And 2-Thiouracil
H
O
NN
N
N
N
H
H
Guanine
+
+
-
2-Thiouracil forms only one hydrogen bond with guanine which is not enough to form a stable pair in the environment of the ribosome A site
+2 Thio-
uracil
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NO
H
NO
N
NH C
ytosine
O
NN
N
N
H
H
Inosine -
+
-
+
-
Wobble Base PairingWobble Base PairingInosine And CytosineInosine And Cytosine
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O
NN
N
N
H
H
Inosine
-
+ NO N
O
NHUracil
+
-
Wobble Base PairingWobble Base PairingInosine And UracilInosine And Uracil
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N
NN
N
H
NH
-
+ Adenine
O
NN
N
N
H
H
Inosine +
-
Wobble Base PairingWobble Base PairingInosine And AdenineInosine And Adenine
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The Wacky Rules ofThe Wacky Rules ofWobble Base PairingWobble Base Pairing
Third codon base:
---------- A or G
----- A
---------- G
---------- U
---------- C or U
---------- C U or G
First anticodon base:
U2-S-UCAGI
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Wobbling and tRNA NumbersWobbling and tRNA Numbers The net effect of wobble base pairing is to reduce the
number of tRNAs that must be produced by a cell In reality cells do not make 61 different tRNAs, one for
each codon Many tRNAs have anticodons that anneal to several
different codons\ Codons are known for which there are more than one
tRNA, although each tRNA carries the same amino acid (i.e., methionine)
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