Molecular Bio, First Post Midterm

Front 1

4 differences between prokaryote and eukaryote transcription

Back 1

1) transcription machinery and DNA sites
2) movement of RNAs from nucleus to different sites for translation
3) post transcriptional modification of mRNA - 5' capping and 3' adenylation
4) exon splicing

Front 2

RNA polymerase II makes what?

Back 2

mRNA, snRNAs, mIRNAs

Front 3

Although additional subunits are in eukaryotic RNA polymerase complex, is eukaryotic RNA similar to prokaryotic RNA?

Back 3

Yes, many components are conserved

Front 4

TATA box?

Back 4

eukaryotic promoter

Front 5

TATA box location?

Back 5

about -35 to -25 upstream of start site

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Inr?

Back 6

initiator sequence around RNA start site

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DPE? location?

Back 7

downstream promoter element, around +30 from start site

Front 8

what does RNA Polymerase II use to identify start site?

Back 8

TATA, Inr, and DPE

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BRE?

Back 9

a DNA sequence found in the promoter region of most genes in eukaryotes and Archaea.[1] [2] The BRE is a cis-regulatory element that is found immediately upstream of the TATA box, and consists of 7 nucleotides.

Front 10

general transcription factors (TFs)?

Back 10

things used by eukaryotes to recognize the TATA box and begin transcription

Front 11

TBP?

Back 11

TATA box binding protein

Front 12

TBP is a subunit of?

Back 12

TFIID

Front 13

Describe TBP

Back 13

highly conserved protein that contains many beta sheets that fit in the minor groove of the TATA box, bending DNA structure

Front 14

In prokaryotes, does RNA polymerase bend DNA?

Back 14

yes

Front 15

After binding of TBP, how does the DNA structure change?

Back 15

It changes to allow TFIIB to bind to BRE upstream of the TATA box

Front 16

TFIIF/RNA Polymerase II complex?

Back 16

complex that binds to the start site

Front 17

TFIIE and TFIIH?

Back 17

Bind to the TFIIF/RNA Polymerase II complex .

Front 18

Preinitiation complex?

Back 18

RNA Poymerase II and all the needed bound TFs

Front 19

when does RNA polymerase II become active?

Back 19

after the preinitiation complex is formed

Front 20

mediator complex?

Back 20

large, 20+ subunit protein that bridges the general transcriptional factors with other regulatory sequences upstream of the TATA box

Front 21

TFIIH role?

Back 21

important subunit of the complex b/c it has a helicase that unwinds DNA for RNA polymerase II

kinase activity that phosphorylates CTD of RNA polymerase II. this kinase activates the RNA polymerase II.

Front 22

CTD?

Back 22

carboxy terminal domain

Front 23

What happens after TFIIH phosphorylizes the CTD of RNA polymerase II?

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Causes RNA polymerase II to disassociate from many of the general transcription factors and allows it to advance beyond the TATA box

Front 24

What is in the CTD: types of amino acids?

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Lots of polar amino acids such as tyrosine, serine, and threonine

Front 25

role of phosphorylated CTD?

Back 25

assembly point for enzymes involved in post transcriptional RNA processing. For ex: the fifth serine is the attachment for the 5' capping enzyme and Ser2 is the attachment for the splicing apparatus

Front 26

How are the two ends of mRNA protected?

Back 26

5' capping and 3' polyadenylation

Front 27

Draw the figure for RNA processing. Show initiation, elongation, capping enzyme, capping, splicing, splicing factors, CBC, spliced intron, polyadenylation factors, termination, poly A tail

Back 27

see figure

Front 28

guanylyltransferase?

Back 28

enzymes that adds a methylated guanine nucleotide to the 5' end of growing mRNA

Front 29

5' cap?

Back 29

modified guanine with methyl

Front 30

CBC?

Back 30

cap binding complex. binds to the mRNA after the 5' cap is put on by the guanylyltransferase

Front 31

What happens after capping the mRNA?

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Ser5 of the CTD is dephosphorylated and the guanylyltransferase disassociates

Front 32

Where is the guanine nucleotide methylated?

Back 32

7' carbon

Front 33

Why does 5' cap have an unusual structure?

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Prevents recognition by RNAases which digest RNAs from the 5' or 3' end

Front 34

Does 5' cap serve as a signal for ribosomes to start translation?

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yes

Front 35

PAP?

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poly A polymerase adds a string of 80-250 adenines to a poly A addition site hard coded in each mRNA transcript

Front 36

poly A tail?

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lots of adenines at the end

Front 37

PABP?

Back 37

poly A binding protein protects the poly A tail from degradation

Front 38

how does endonuclease work in 3' polyadenylation?

Back 38

endonuclease cleaves the mRNA at the poly A addition site and leaves two stumps. the 5' side of the stump is the nascent mRNA and will have adenines added to it by the PAP

Front 39

Draw the 3' polyadenylation.

Back 39

draw the card

Front 40

In eukrayotes, what is analogous to prokaryote rho termination?

Back 40

termination of transcription, torpedo model

Front 41

Draw the torpedo model

Back 41

notes

Front 42

what happens in torpedo model?

Back 42

mRNA with a poly A tail is released from RNA polymerase II and a "stump" of mRNA is left hanging and this stump is unprotected without a 5' cap. The free 5' end is then attacked by RNAase. The RNAase moves faster than the RNA polymerase II and the RNAase induces the RNA polymerase II to disassociate from DNA

Front 43

oligo-DT?

Back 43

primes reverse transcriptase in RT-PCR. allows us to selectively isolate mRNA as opposted to tRNA or rRNA for reverse transcription and amplification

Front 44

draw the function of oligo-DT

Back 44

notes

Front 45

the enzymes that mediate 5' capping and 3' polyadenylation are closely tied to what?

Back 45

CTD of RNA polymerase II

Front 46

are introns always junk DNA?

Back 46

no

Front 47

what info do introns contain?

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contain info about how exons should be spliced together.

they also encode noncoding RNA molecules such as microRNAs which are important regulatory elements

Front 48

alternative splicing?

Back 48

different proteins come from the same gene

Front 49

Are the 5' and 3' junctions splice sites?

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yes

Front 50

branch points?

Back 50

splice sites inside introns

Front 51

describe rna splicing with a branch point, 3 steps

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1) branch point adenine attacks teh 5' splice site with Oxygen
2) freed 5' splice site attacks teh 3' splice site
3) exons are spliced togther and intron is released as a lariat

Front 52

lariat?

Back 52

freed intron from rna splicing

Front 53

spliceosome?

Back 53

massive catalytic complex with protein and RNA. it's where RNA splicing happens.

Front 54

snRNAs?

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small nuclear RNA

Front 55

snRNP?

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small nuclear ribonucleoprotein. bring various sites of intron together to catalyze splicing reactions.

Front 56

because the spliceosome contains RNA, can various sites of the intron be recognized?

Back 56

yes

Front 57

where is the info for how RNA should be transcribed carried?

Back 57

introns and exons

Front 58

what catalyzes the excision of introns involving 2 sequential nucleotide attacks?

Back 58

snRNPs

Front 59

what catalyzes peptide bond formation in euakryotes and prokaryote?

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eukaryotes: 28S and 5.8S
prokaryotes: 23S

Front 60

what catalyzes recognition of translation start site in eukaryotes and prokaryotes?

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18S and 16S

Front 61

eIF1?

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eukaryotic initiation factor 1. occupies the E site.

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eIF1A?

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eukaryotic initiation factor 1A. occupies the A site.

Front 63

eIF3?

Back 63

blocks subunit assembly.

Front 64

what is analogous to prokaryote IF1 and IF3?

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prokaryote IF1 : eukaryote eIF1A
prokaryote IF3: eukaryote eIF1

Front 65

tRNA _ i_ Met?

Back 65

special tRNA used for translation initiation in eukaryotes

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eIF2?

Back 66

GTPase. it brings initiator tRNA to the P site

Front 67

eIF5B?

Back 67

GTPase. it joins with eIF1A.

Front 68

43S initiation complex?

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small ribosome, eIF1, Met-tRNA_i_Met, eIF2 with GTP, eIF1A, eIF5B with GTP, eIF3

Front 69

How is mRNA marked for translation?

Back 69

5' cap and 3' poly A tail

Front 70

eIF4F?

Back 70

complex that recognizes 5'cap and 3'poly A tail. delivers mRNA to the 43S initiation complex

Front 71

kozak sequence?

Back 71

designates start codon in eukaryotes.

Front 72

how is kozak diff from shine dalgarno?

Back 72

kozak includes start codon and requires a purine 3 bp before the start codon

Front 73

can an assembled 43S complex scan along mRNA and look for kozak sequence?

Back 73

yes

Front 74

what happens after kozak sequence is found?

Back 74

eIF5B hydrolyzes GTP to release all initiation factors. 60S subunit joins 40S subunit to start translation.

Front 75

what 2 things have to be recognized for translation to start?

Back 75

kozak sequence and cap

Front 76

what is analogous to prokaryotic EF-Tu, EF-Ts, and EF-G?

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Ef-Tu: eEF1alpha
Ef-Ts: eEF1beta gamma
EfG: eEF2

Front 77

does RRF exist in prokaryotes?

Back 77

no

Front 78

eRF1?

Back 78

single release factor corresponds to prokaryote RF1 and RF2

Front 79

eRF3?

Back 79

corresponds to RF3

Front 80

ABCE1?

Back 80

protein that is kind of like RRF

Front 81

can eIF1, 1A, and 3 dissociate into 2 subunits?

Back 81

yes

Front 82

are there lots of differences between prokaryotic and eukaryotic initiation?

Back 82

yes

Front 83

are the proteins involved in prokaryotic and eukaryotic elongation and termination similar?

Back 83

yes

Front 84

Know the table 18-2

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notes

Front 85

first tRNA in eukaryotes and prokaryotes

Back 85

eukaryotes: Methionyl-tRNA (met)

prokaryotes: N-Formylmethionyl - tRNA (fmet)