Hebrews 1:1-2

©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;


TranscriptionTranscriptionFrom DNA To RNAFrom DNA To RNA

Timothy G. Standish, Ph. D.

DNA

mRNA

Transcription

IntroductionIntroduction

The Central Dogma The Central Dogma of Molecular Biologyof Molecular Biology

Cell

Polypeptide(protein)

TranslationRibosome


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


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


Stages of TranscriptionStages of Transcription

1Initiation

2Elongation

3Termination


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


Prokaryotic Transcription InitiationProkaryotic Transcription Initiation

Constitutive GeneHeat Shock GeneP1 P2

Different promoters

RNAPol.


Prokaryotic Transcription InitiationProkaryotic Transcription Initiation

Constitutive GeneHeat Shock Gene P2P1

Different promoters

RNAPol.


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


RNA Pol.

InitiationInitiation

Promoter

RNA Pol.T. F.

T. F.

T. F.

5’RNA


RNA Pol.

InitiationInitiation

RNA Pol.

T. F.

5’RNA

Promoter

T. F.

T. F.


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


RNAPol.

5’RNA

RNA Pol.

5’RNA

TerminationTerminationRho IndependentRho Independent

Terminator


RNAPol.

5’RNA RNA

Pol.

5’RNA

TerminationTerminationRho IndependentRho Independent

Terminator


RNAPol.

5’RNA

TerminationTerminationRho DependentRho Dependent

Terminator

RNA Pol.

5’RNA The terminator

sequence slows RNA polymerase


RNAPol.

5’RNA


Terminator

Help, Rhohit me!

RNA Pol.

5’RNA

Rho catches up with RNA polymerase


RNAPol.

5’RNA


Terminator

RNA Pol.

5’RNA

The elongation complex disintegrates


DifferencesDifferencesBetween Transcription InBetween Transcription In

Prokaryotes and Prokaryotes and EukaryotesEukaryotes


3’

5’

5’

3’

Transcription And Translation Transcription And Translation In ProkaryotesIn Prokaryotes

Ribosome

Ribosome5’

mRNA

RNAPol.


DNA

Cytoplasm

Nucleus

Eukaryotic TranscriptionEukaryotic Transcription

ExportG AAAAAA

RNA

Transcription

Nuclear pores

G AAAAAA

RNAProcessing

mRNA


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


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


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


Common Splicing MechanismCommon Splicing MechanismLariat Lariat FormationFormation

Exon 2A AG

GU

3’

5’

3’

Exon 1Intron

Lariat

Yee ha!Lariat



3’

Exon 2Exon 15’

A

AG

GU

Intron lariat

Following excision, the lariat is rapidly degraded



3’

Exon 2Exon 15’

Following excision, the lariat is rapidly degraded


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


Wobble Base Wobble Base PairingPairing


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


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


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


Some Other Strange tRNA BasesSome Other Strange tRNA Bases

NO

H

NO

N

NH C

ytosine

H

O

NN

N

N

N

H

H

Guanine -+

+

+

-

-

Base PairingBase PairingGuanine And CytosineGuanine And Cytosine

N

O

N

ON

H+

- UracilN

NN

N

HN H

-

+Adenine

Base PairingBase PairingAdenine And UracilAdenine And Uracil

Base PairingBase PairingAdenine And CytosineAdenine And Cytosine

NO

H

NO

N

NH C

ytosine-

+

-

N

NN

N

HN

H

-

+

Adenine

N

O

N

ON

H+

- Uracil

Base PairingBase PairingGuanine And UracilGuanine And Uracil

H

O

NN

N

N

N

H

H

Guanine

+

+

-

NO N

O

NHUracil

+

-

Wobble Base PairingWobble Base PairingGuanine And UracilGuanine And Uracil

H

O

NN

N

N

N

H

H

Guanine

+

+

-

N

S

N

ON

H+

-2 Thio-

uracilN

NN

N

HN H

-

+Adenine

Base PairingBase PairingAdenine And 2-ThiouracilAdenine And 2-Thiouracil

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

NO

H

NO

N

NH C

ytosine

O

NN

N

N

H

H

Inosine -

+

-

+

-

Wobble Base PairingWobble Base PairingInosine And CytosineInosine And Cytosine

O

NN

N

N

H

H

Inosine

-

+ NO N

O

NHUracil

+

-

Wobble Base PairingWobble Base PairingInosine And UracilInosine And Uracil

N

NN

N

H

NH

-

+ Adenine

O

NN

N

N

H

H

Inosine +

-

Wobble Base PairingWobble Base PairingInosine And AdenineInosine And Adenine


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


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)

Hebrews 1:1-2

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Transcript of Hebrews 1:1-2