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Ch. 18,21 : Gene Expression: The Genetic Code & Transcription


Gene Expressi On
  
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Gene expression -central dogma of molecular bio:
 
the use of info in DNA to direct the production of proteins -DNA-RNA-protein
Nuclear env protects
 
chromo and immature RNA from cytoplasmic enzymes
Nuclear pores
 
direct continuity between cytoplasm & nucleoplasm -mammalian nucleus ~3,000-4000 pores -eukaryotic cells must transport macrmolecues in & out of nucleus
Where does replication and trancrption occur? and where does translation occur?
 
-replication and transcription in nucleus, translation in cytosol
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)
Transcription occurs in the
 
Nucleus (eukaryotes) Cytosol (prokaryotes
A cell uses 3 kinds of RNA to make proteins
 
messenger RNA (mRNA) transfer RNA (tRNA) ribosomal RNA (rRNA)
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
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
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
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
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
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
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
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
Elongation (euk.)
 
need special proteins to dissassemble nucleosome in front of RNA poly & reassemble behind RNA poly
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
Ribosomal RNA
 
-the most abundant (70-80%), most stable RNA in cells- component of ribosomes
Transfer RNA
 
- 10-20%- critical in protein syn
Messenger RNA
 
-less than 10%- amino acid sequence
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
Methylation
 
addition of methyl groups-protect from enzyme degredation (esp rRNA) then transport to cytosol for translation
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
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
Introns
 
-intervening sequences within the euk primary transcript absent in mature functional RNA- not expressed -can be 99% of genes DNA
RNA spliced...
 
removes introns, joins exons (expressed sequences) -by ribozymes- self-splicing RNA introns -by spliceosomes- RNA-protein complex
Catalyzed by spliceosome -
 
RNA & protein complex (small nuclear ribonucleoproteins, snRNPs) -recognizes 5' to 3' splice sites-cut & link exons
Why do eukaryotic genes have introns? 1 of 5
 
-generate numerous diff mRNA's from 1 RNA transcript through alternative RNA-splicing-exon shuffling
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
3 of 5
 
Evolutionary- hasten evolution of new and potentially useful proteins
4 of 5
 
each exon codes for diff fct'l regions- protein can fold independently into separate domain
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
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
(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
Exceptions- few slight differences (translation)
 
-In mitochondria, a few bact & unicellular org's -UGA-21st amino acid selenocysteine, UAG-22nd amindo acid-pyrrolysine

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