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Biochemistry > RNA And Protein Synthesis > Flashcards

RNA And Protein Synthesis Flashcards

1
Q

Ribosomal RNA

A 
. 4 of them (28S, 18S, 5.8S, and 5S)
. Assoc. w/ proteins to form ribosomes 
. Required for protein synthesis 
. Accounts for 80% total RNA in cell 
. Non-coding (functional)
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2
Q

Transfer RNA

A 
. Carry AA
. 1 specific tRNA for each AA 
. Required for protein synthesis 
. Make up 15% total RNA 
. Non-coding (functional)
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3
Q

Messenger RNA

A

. Carry genetic info encoded by DN from nucleus to cytosol
. Transcripts (protein coding)
. Heterogenous (size and sequence differs)
. 5% total RNA

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4
Q

Long non-coding RNA (lncRNA)

A

. Antisense: transcribed from sense strand of gene nucleotide sequence, not processed/no splicing, bind mRNAs transcribed from antisense strand to repress gene expression
. Sense: transcribed and processed like primary transcripts of protein-coding genes, regulate neighboring genes
. Non-coding (functional)

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5
Q

Micro RNA (miRNA)

A

. 22 nucleotides in length
. Bind to 3’ UTR of specific mRNAs
. Regulate gene expression by repressing protein production/triggers mRNA degradation
. Non-coding (functional)

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6
Q

MiRNAs in human disease

A

. RNA interference (gene regulation via miRNA)
. Critical for normal development
. 50% protein-coding genes regulates by this
. A;teres patterns of miRNA shown to be assoc. w/ various diseases and cancers

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7
Q

Protein-coding gene

A

. Genes w/ nucleotide sequence that is transcribed and is translated to produce protein
. Divided into coding and regulatory region/transcriptional start site
. Coding region divided into coding Exons and non-coding introns

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8
Q

Consensus sequences

A

. Recognition markers/landmarks for protein binding
. DNA sequences highly conserved and found in noncoding regions
. Regions that modulate gene expression
. Bound by transcription factors or other regulatory proteins
. Ex: TATA box

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9
Q

Promoter sequence

A

. Region that selects or determines start site of RNA synthesis
. Contains TATAbox 15-30 bps upstream from transcription start site

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10
Q

Binding of transcription factors to TATA box facilitates recruitment of _____

A

RNA polymerase II

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11
Q

Splice acceptor and donor sequences

A

. Consensus sequences found at 5’ and 3’ ends of introns
. Needed for removal of introns out of primary transcript (splicing)
. Introns start w/ splice donor site (always GU, GT in DNA) and end w/ splice acceptor site (Ends w/ AG)

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12
Q

Transcription

A

. Production of RNA
. Occurs at nucleus
. only 1 of 2 strands is used as template (template strand)
. Catalyzed by RNA polymerase that read DNA in 3’-5’ direction to produce complementary RNA, have proofreading activity

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13
Q

RNA polymerase I

A

Synthesized precursors of rRNA in nucleolus

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14
Q

RNA polymerase II

A

. Produces mRNAs and miRNAs in nucleus

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15
Q

RNA polymerase III

A

. Catalyzes synthesis of tRNAs and 5S rRNAs in nucleus

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16
Q

Steps in production of mRNA by RNA polymerase II (RNAP)

A

. Initiation: chromatin remodeling (to open), assembly of preinitiation complex and recruitment of RNAP
. Elongation: local unwinding of DNA, RNAP moves along template strands catalyzes creation of transcript that grows in 5’-3’
. Termination: RNAP encounters stop signal, enzyme and transcript released

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17
Q

Preinitiation complex

A

. 6 general transcription factors (GTFs)
. Each GTf is a multiprotein complex
. Proper assembly required for proper recruitment and correct positioning of RNAP

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18
Q

Typical structure of mRNA

A

. Produced in cytosol
. Have exons and introns
. When initially produced, contain regulatory sequences at 5’ and 3’ ends called untranslated regions (needed for trailing and translational efficiency) and are not part of product

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19
Q

Regulatory features of mRNA transcription through RNAP II

A

. Region upstream of start site are regulatory

. Basal promoters, enhancers/repressors, and response elements

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20
Q

Basal Promoters

A

. Ensures basal expression
. Found adjacent to transcriptional start site
. TATA box: prox. Element, specifics start site by directing RNAP to correct site, ensures fidelity of initiation
. CAAT and GC boxes: protein-DNA interactions determine frequency of transcription initiation

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21
Q

Enhancers/repressors, insulators, and response elements

A

. Participate in regulated expression
. Found far away from start site
. Enhance/repress expression
. Mediate response to signals/hormones by inc/dec. rate of initiation
. Function in orientation-independent manner

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22
Q

RNA transcription w/ introns is called ____

A

. Primary transcript
. Pre-mRNA
. Heterogenous nuclear RNA (hnRNA)
.

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23
Q

How pre-mRNA is processed to get to cytosol

A

. Addition of 5’ cap (7-methylguanosine triphosphate) to 5’ end of transcript
. Addition of poly A tail: signaled by polyadenylation signal (AAUAAA) at 3’ end of primary transcript
. Splicing of introns: removed, needs spliceosome

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24
Q

Spliceosome

A

. Combo of pre-mRNA and complex of 5 small nuclear RNAs (U1, 2, 5, and 4/6) and over 50 proteins (snRNPS)

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25
Q

Branch site in intron splicing

A

. Hydroxyl group of adenosine in intron required for splicing

26
Q

Exonic splicing enhancers and silencers (ESE and ESS)

A

. Influence constitutive and alternative splicing

. Commonly altered in human disease

27
Q

Steps of intron splicing

A

. 1: spliceosome recognizes splice donor/acceptor site
. 2: branch site OH attacks 5’-phosphate of splice donor at 5’ end of intron to form 2’-5’ phosphodiester forming lariat when cleaved
. 3: 3’-OH of upstream exon attacks splice acceptor at 5’-P of downstream exon, intron removed as phosphodiester bond forms to join exons
. Exported into cytoplasm through nuclear pores

28
Q

Rifampin uses and mechanism

A

. Antibiotic used to treat TB
. Prevents bacterial cell growth by inhibiting RNA synthesis
. Targets bacterial DNA-dependent RNA polymerase enzyme
. Binds to beta-subunit of bacterial RNAP to block transcription initiation

29
Q

Dactinomycin (Actinomycin D)

A 
. Antibiotic/anti-neoplasticism agent used to treat Wilm’s tumor and sarcomeres
. Prevents bacterial growth by inhibiting RNAP movement and RNA elongation during DNA-primed RNA synthesis by binding to DNA template
. Member of actinomycine class of antibiotics isolated from soil bacteria of Strep
30
Q

Alpha-amanitin

A

. Toxin in mushrooms
. Inhibits human RNAP II movement along DNA template to prevent RNA synthesis
. Symptoms after 24 hrs (diarrhea and cramps) then liver and kidney damage preceding total system failure and death w/in 1 week

31
Q

Codons

A

. Sequence of 3 nucleotides for specific amino acid
. Written and read in 5’-3’ direction
. 64 different ones
. 61 of 64 code from 20 common AA
. 3 are termination signals (UAG, UGA, and UAA)

32
Q

Characteristics fo genetic code

A

. Specific: codon always codes for same AA
. Universal: genetic code conserved across species
. Degenerate/redundant: AA may be specified by more than one codon
. Nonoverlapping and commaless: code read continuously from fixed point, sequence of bases processed 3 at a time

33
Q

Components required for translation

A

. MRNA: template
. AA: must be available to put in final product
. TRNA: translators of genetic code
. Ribosomes: rRNA and protein complex
. Protein factors: specific initiation, elongation, and termination factors required
. Energy sources: ATP and GTP

34
Q

TRNA structure

A

. AA attachment site at 3’ end

. Anticodon, 3-base recognition requests that allows complementary binding to correct series of 3 bases on mRNA

35
Q

Aminoacyl-tRNA synthetases

A

. Highly selective enzymes
. Needed for attachment of AA to corresponding tRNAs to form aminoacyltRNAs
. Uses ATP to catalyze 2-step rxn resulting in covalent attachment of carboxyl group of AA to 3’ end of tRNA

36
Q

Ribosome mechanism

A

. 60S subunit catalyzes formation of peptide bonds btw AA
. 40S subunit binds mRNA, responsible for accuracy of translation
. A, P, and E binding sites for tRNA
. A: binds incoming aminoacyl-tRNA, specifies next AA added to polypeptide chain
. P: peptidyl-tRNA carries growing chain
. E: empty tRNA that is about to exit ribosome

37
Q

How tRNA is attached to it’s specified AA

A

. Covalently attachment
. Charged when bound to AA, uncharged when not bound to AA
. When AA is attached to tRNA it is activated

38
Q

Post-translational modification of protein types

A

. Co-translational or post-translational

39
Q

Proteolytic cleavage (trimming)

A

. Most common
. Proteolysis removal of their leading Met residue after it emerges from ribosome
. If proteins synthesized as inactive precursors, they are activate by limited proteolysis (trypsinogen) or removal of polypeptides (proinsulin to insulin)

40
Q

Covalent posttranslational modifications

A

. Modified by covalent attachment to chemical groups, sugars, lipids, or small proteins to specific AA

41
Q

Hydroxylation posttranslational modifications

A

. Pro and Lys residues of collagen alpha-chains hydroxylated in ER
. Requires Vit. C

42
Q

Phosphorylation as posttranslational modification

A

. Hydroxyl groups of Ser and Thr and sometimes TYr residues
. Catalyzed by protein kinases, reversed by phosphatases
. Activates or inhibits function

43
Q

Glycosylation as posttranslational modification

A

. Attachment of carb chains to hydroxyl groups of Ser or Thr (O-linked) or Asn (N-linked)
. Normally secreted from cell or become part of plasma membrane

44
Q

What does the 5’ cap of mRNA do?

A

. Prevents degradation by exonuclease

. Facilitates binding of mRNA to ribosomes during protein synthesis

45
Q

How is it added and What does addition of poly(A) tail to mRNA do?

A

. Signal recognized by endonuclease that cleaves RNA about 20 nucleases
. New 3’ end serves as primer for enzymatic addition of 250 adenine ribonucleotides
. Important for nuclear export, translation, and mRNA stability

46
Q

Premature termination codons

A

. splicing mutation creates truncated protein w/ impaired function

47
Q

Nonsense mediated decay

A

. Doesn’t act on single exon genes, depends on RNA splicing
. Exon-junction complexes bind 20 nt upstream of 3’ end of transcribed exons and remain bound in mature mRNA
. Ribosome removes them in translation one-by-one as protein synthesis progresses
. If PTC detected, it is released from ribosome w/ EJCs bound to signal degradation

48
Q

Where will fame-shift PTC occur and escape NMD most likely?

A

55 nt upstream of last exon

49
Q

Sonic hedgehog molecule

A

. Secreted signaling molecule
. Critical role during CNS development, face, limbs, and other body parts
. Expressed and secreted to form morphogenetic gradient that influences tissue patterning
. Coding sequence mutations are a cause of holoprosencephaly (HPE)

50
Q

. Zone of polarizing activity in vertebrate limb importance

A

. Specific expreassign of SHH here is critical to having normal limbs
. ZPA Regulatory sequence (ZRS): long-distance enhancer element identified as direct SHH expression to ZPA

51
Q

ZRS enhancer sequence is found ____

A

. 1st intron of neighboring LMBR1 gene
. 1 Mb away from SHH
. Mutations cause polydactyly

52
Q

ZRS enhancer point mutations cause ___

A

Preaxial polydactyly (double thumb)

53
Q

Position effects

A

Chromosomal rearrangements caused by repositioning of long range enhancers required for normal gene expression
. Creates mutations like the thalassemias

54
Q

Imitating codon for translation

A

AUG, codes for methionine

55
Q

Rate limiting step of translation

A

. Binding of methylguanosine cap binding complex to the 5’ cap of mRNA

56
Q

elongation in translation steps

A

. Binding of aminoacyl-tRNA to the A site (eukaryotic elongation factors assist in delivery of aminoacyl-tRNA whose codon appears next)
. Peptide bond formation: alpha-amino group of new aminoacyl-tRNA in A site carries nucleophilic attack on esterified carboxyl group of peptidyl-tRNA occupying P site
. Rxn catalyzed by peptides-transferase activity of 60S ribosomal subunit
. Growing peptide chain now attached to tRNA in A site

57
Q

Translocation steps in translation

A

. Peptide moiety is removed from tRNA in P site
. TRNA dissociates from P site and moves into E site
. Elongation factors tranlocate the newly formed peptidyl-tRNA into empty P site and A site becomes free

58
Q

How primary miRNAs (pri-miRNAs) transcribed from genome?

A

. Via RNA Pol II or III
. Shorter inverted sequences that form double stranded RNA stem loop structure
. Processed into pre-miRNAs stemloop in nucleus by microprocessor complex comprised of DGCR8 and drosha (ribonuclease)

59
Q

How pre-miRNAs is modified before translation?

A

. Pre-miRNA exported into cytoplasm via exportin 5/Ran GTP complex
. Further processed into mature miRNA by Dicer
. 1 of 2 strands of mature miRNA is loaded into 1 of 4 members of argonaute family to form miRISC complex

60
Q

How miRNA is translated

A

. Chosen strand is guide miRNA
. MiRISC complex binds mRNAs at miRNA response elements (MREs)
. If guide miRNA contains mismatch in nucleotide sequence, gene expression may be inhibited through mechanisms leading to mRNA degradation or translational repression
. If guide RNA in miRISC complex is perfect match to an mRNA’s MRE, then RNA endonuclease activity cleaves mRNA

Biochemistry (64 decks)

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