Ch. 18,21 : Gene Expression: The genetic code & Transcription

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1. 

Gene expression

-central dogma of molecular bio:

 

the use of info in DNA to direct the production of proteins

-DNA-RNA-protein

 
2. 

Nuclear env protects

 

chromo and immature RNA from cytoplasmic enzymes

 
3. 
Nuclear pores
 

direct continuity between cytoplasm & nucleoplasm

-mammalian nucleus ~3,000-4000 pores

-eukaryotic cells must transport macrmolecues in & out of nucleus

 
4. 
Where does replication and trancrption occur? and where does translation occur?
 

-replication and transcription in nucleus, translation in cytosol

 
5. 
Nuclear pores
 

-Inner & outer membranes nuclear env fuse at nuc pore complex with intricate protein structure

-small molecules (<10nm) diffuse through nuc pores (dNTP's, ions, small proteins-histones)

 
6. 
Transcription occurs in the
 

Nucleus (eukaryotes)

Cytosol (prokaryotes

 
7. 
A cell uses 3 kinds of RNA to make proteins
 

messenger RNA (mRNA)

transfer RNA (tRNA)

ribosomal RNA (rRNA)

 
8. 
Overview of Transcription:
 

Binding of RNA polymerase to DNA promoter sequence

Initiation of RNA synthesis- base pair w/DNA template

Elongation of RNA strand

Termination at termination signal, release RNA molecule, RNA polymerase dissociates from DNA molecule

 
9. 
Binding- RNA polymerase binds at DNA promotor site
 

-the transcriber is RNA polymerase

-consists of 2 alpha, beta and beta prime and sigma factor

- complete holoenzyme- ^ required to ensure initiaiton at proper DNA site

-Sigma factor ensures correct binding of RNA poly to promotor

 
10. 
Promoter
 

is upstream (neg #), before transcribed sequence/gene- downstream (pos #), no # "0"

-specific sequence, determines which strand to serve as template

-statr point (+1) described 5'-3' on coding strand (opp template strand)

-not identical, have consensus sequences (most common)

-mutations interfere w/promoter fct/activity

-triggers local unwinding of DNA strands by topoisomerase

 
11. 

INITIATION

 

-link up dNTP's in 5'-3' direction to make RNA strand

-RNA-DNA hybrid- complementary base pairing to DNA template

-Add until ~9 nucleotides long

 
12. 
ELONGATION
 

-RNA poly core enzyme (2 alpha,1 beta,1beta prime, prime) reads along the DNA, untwist DNA double helix

-Adds nucleotides to RNA strand

-Proofreading- RNA poly has 3'-5' exonuclease activity-correct mistakes-remove wrong nucleotides,insert correct bases

 
13. 

TERMINATION- reach termination signal

type 1 of 2:

 

1-Need rho (p) factor

-binds to specific 50-90 base termination sequence

-needs ATP to unwind & release new RNA from DNA template

-Release RNA poly

: RNA poly can bind sigma factor, start another RNA synthesis event

 
14. 
type 2 of 2:
 

2-Does not need rho (p) factor

-RNA has short complementary sequence to each other & short GC-rich sequence, w/several U's

-RNA forms hairpin loop

-pulls RNA away from DNA

: AU bonds- weaker

 
15. 

Differences in Eukaryotic Transcription

Protein-protein interactions

 

-Need transcription factors (TF)- must bind to DNA before RNA poly can bind to promoter

-Bind to other proteins & DNA

-Recruit coactivator proteins- help assymbly of RNA poly complex to initiate RNA syn

-Unwinds DNA

 
16. 
Elongation (euk.)
 
need special proteins to dissassemble nucleosome in front of RNA poly & reassemble behind RNA poly
 
17. 
Termination (Euk)
 

-many signals depending on type of RNA poly

-short run of U's

-RNA cleavage:

special AAUAAA signal - cut 10-13 bases downstream, then add poly(A) tail at cleavage site

-RNA processing

 
18. 
Ribosomal RNA
 
-the most abundant (70-80%), most stable RNA in cells- component of ribosomes
 
19. 
Transfer RNA
 
- 10-20%- critical in protein syn
 
20. 
Messenger RNA
 
-less than 10%- amino acid sequence
 
21. 
rRNA
 

-single primary transcript unit with transcribed spacers

-150-200 copies in cell

-Methylation of ribose sugars by small nucleolar RNA's (snoRNA's)

-series of cleavage reactions to release mature rRNAs

-transcribed spacers are degraded

-chemically modify euk primary transcript into mature RNA

 
22. 
Methylation
 
addition of methyl groups-protect from enzyme degredation (esp rRNA) then transport to cytosol for translation
 
23. 
5' methyl guanine cap
 

to 5' end of pre-mRNA & mRNA

-added after initiation of RNA syn

-makes RNA more stable so cannot be degraded

-position of ribosome to start translation

 
24. 
Poly (A) tail
 

50-250 nucleotides of A's added to 3' tail

-NOT FROM DNA SEQUENCE-found in termination signal

-added by poly (A) polymerase after special AAUAAA signal

-make it more stable, longer life span

-help export mRNA from nucleus to cytoplasm

-help ribosome recognize mRNA to be translated

 
25. 
Introns
 

-intervening sequences within the euk primary transcript absent in mature functional RNA- not expressed

-can be 99% of genes DNA

 
26. 
RNA spliced...
 

removes introns, joins exons (expressed sequences)

-by ribozymes- self-splicing RNA introns

-by spliceosomes- RNA-protein complex

 
27. 
Catalyzed by spliceosome -
 

RNA & protein complex (small nuclear ribonucleoproteins, snRNPs)

-recognizes 5' to 3' splice sites-cut & link exons

 
28. 

Why do eukaryotic genes have introns?

1 of 5

 

-generate numerous diff mRNA's from 1 RNA transcript through alternative RNA-splicing-exon shuffling

 
29. 
2 of 5
 

-help increase biological complexity increase in gene number (25,000 genes produce 200,000-1 million proteins

-ex: immune system

-bacteric and archaea lost introns from their streamlined genomes

 
30. 
3 of 5
 
Evolutionary- hasten evolution of new and potentially useful proteins
 
31. 
4 of 5
 
each exon codes for diff fct'l regions- protein can fold independently into separate domain
 
32. 
5 of 5
 

Nucleic acid editing- more 1-100's nucleotides inserted or converted

-can inactivate retroviruses

-ex. HIV has defense protein- destroys DNA-editing enzyme

 
33. 
Translation
 

converts nucleotides of a gene into amino acids in a protein in order using genetic code

-almost universal- prokaryotes, euk, & viruses

-established early on in history of life on Earth

 
34. 
(translation) mRNAs are...
 

"read" by ribosome in 3-nucleotide units, termed

(64 possible combinations)

-each codon codes for an amino acid or stop signal

-code is degenerate- adaptibility, some are silent mutations

 
35. 
Exceptions- few slight differences (translation)
 

-In mitochondria, a few bact & unicellular org's

-UGA-21st amino acid selenocysteine, UAG-22nd amindo acid-pyrrolysine