Chapter 11:DNA Transcription and Translation Flashcards
Where is a segment of DNA transcribed?
In the nucleus
Where is mRNA translated?
cytoplasm
As more mutations occur in DNA what risk increases?
cancer
Does translation always have to be perfect?
NO but transcription within the nucleus does
List characteristics of DNA!
DNA:
- T= A
- double stranded
- 250 000 000nt
- DNA poly
- Primer
- 1 in 10^7 nt errors
List haracteristics of RNA!
- U= A
- single stranded
- 1000 nt
- RNA Poly
- no primer
- 1 in 10^4 nt errors
Does RNA poly have more errors than DNA poly?
- YES
- it does not require a poly like DNA but it has many more errors but that is okay because it is not a fixed representation of the DNA
What does it mean that RNA is a mobile nucleic acid?
- permits cell to separate information storage in the nucleus ( or not if you’re a prokaryote) into information utilization which is the RNA
How can you amplify the amount of protein in a cell?
- by transcribing/transcription many times
DNA Transcription:
- Double stranded DNA must be ___ to expose a single stranded DNA template.
- unwound
DNA Transcription:
- Ribonucleotide addition is to the ___ DNA strand.
- non coding DNA strand
DNA Transcription:
- there is a temporary __/__ hybrid?
- DNA/RNA hybrid
~9nt
DNA Transcription:
- With your DNA it is double stranded and there are certain sequences within your DNA that tell the RNA poly ?
- which strand needs to be used as the template and which is not
DNA Transcription:
- What is the template used for?
- mRNA transcription (well RNA)
DNA Transcription:
Which of these strands of DNA is the coding strand and which is the non coding strand?
5’AATTGGC3’
3’TTAACCG5’
- AATTGGC = coding strand
- TTAACCG = non coding strand
DNA Transcription:
After transcription what will the following coding strand’s mRNA strand be?
3’TTAACCG5’
- AAUUGGC
DNA Transcription:
What is a protein?
- amino acid chain (polypeptide)
DNA Transcription:
Protein is coded into ___ via ____ which are a pile of nucleic acids which are arranged via complementary strands which make up genes.
- DNA via chormosmes
DNA Transcription:
- Genes CODE for proteins. Code??
- a certain arrangement of nucleotides give you nucleic acids so the gene is telling you that particular sequence of aa that will give you the resulting protein
DNA Transcription:
- Gene?
- a sequence of nucleotides that tell us the sequence of aa’s for a protein
DNA Transcription:
- why is the sequence of nucleotides in the gene extremely important?
- because if you take the wrong genes you are not getting the correct sequence of aa’s
DNA Transcription:
- coding strand?
- it’s sequence in the gene codes for the correct sequence of amino acids
DNA Transcription:
- non-coding strand?
- the complementary strand to the coding strand..
DNA Transcription:
-RNA is synthesized on?
- a complementary DNA strand ..it needs to choose the correct one so it will be read during translating giving us the right aa’s
DNA Transcription:
- What is the RNA sequence and what is it coming from?
- It needs to be the exact same as the coding strand this is ensured by using the non coding DNA strand as a template! So you are not actually touching the sequence of DNA that codes for the genes that give us the protein
What are the three Eukaryotic RNA polymerases that we discussed in class?
- RNA Poly I
- RNA Poly II
- RNA Poly III
Eukaryotic RNA Polymerases:
- RNA Poly I?
- rRNA - structural (ribosomal RNA which are not translated so they stay in an RNA state and are used along with ribosomes to make proteins.
Eukaryotic RNA Polymerases:
- RNA Poly II?
- mRNAs- informational
Eukaryotic RNA Polymerases:
- RNA Poly III?
- tRNA’s (transfer RNA’s…transfer aa’s), snRNA’s (splicing …getting rid of introns) and ribosomal 5s RNA
Eukaryotic Nuclear RNA Polymerases.
Enzymes? (5)
- RNA Poly I - synthesizes larger rRNAs (28S, 18S and 5.8S)
- RNA Poly II- synthesizes mRNAs, most small nuclear RNAs( snRNAs and snoRNAs) most micro RNAs and telomerase RNA
- RNA Poly III - small RNAs, including tRNAs, 5S rRNA and U6 snRNA
- RNA Poly IV and V (plants only)- siRNAs
Eukaryotic RNA Polymerases:
-rDNA?
- rRNA genes in 100’s of copeies (nucleolus region)
Eukaryotic RNA Polymerases:
- nucleolus region?
- makes subunits and transcribing rRNA so in us when you see those granular regions…RNA genes are in clusters …there are certain regions on each of our chromosomes that look like that and are called this .
Initiation of RNA Synthesis in Bacteria:
- core enzyme?
-RNA poly with no roman numerals
Initiation of RNA Synthesis in Bacteria:
- Sigma factor?
- associates with the core enzyme helping it join to the dsDNA
Describe the Initiation of RNA Synthesis in Bacteria!
- Loose association between DNA and core enzyme. RNA chains that are begun are not initiated at proper sites.
- Complete enzyme (holoenzyme) = Sigma factor + core enzyme)
- Association of complete enzyme with DNA (in promoter region) at proper site and opening of the double helix.
L> proper site= -35 and -10 site that is specific for them to sit on and associate with DNA
Initiation of RNA Synthesis in Bacteria:
- where is the start of synthesis?
+1
Initiation of RNA Synthesis in Bacteria:
- upstream?
- downstream?
- to the left (-35 and -10)
- to the right (+1?)
What are the steps to RNA synthesis in Bacteria?(4)
- Recognition: sigma factor permits the RNA poly to recognize promoter regions in DNA at that time when it comes in once it associates with DNA it will actually unwind DNA at that point aka exposed template after recognition has occurred
- Initiation: warm up occurs… needs to get started…RNA poly… 10-12 nt are synthesized and then released…now RNA poly is ready to go.
- Elongation: RNA poly synthesizes 50nt per second…a hybrid of RNA and DNA is created (ant long)…so 50nt per second are added to the molecule which occurs until termination…once elongation occurs sigma is let go
- Termination: RNA poly will stop and dissociate
Basic Elements of a Promoter Region in E. coli DNA:
-promoter region?
-promotes transcription -> sequence of DNA upstream from the +1 site of the gene itself
Basic Elements of a Promoter Region in E. coli DNA:
- (-35)?
consensus sequence from +1 TTGACA/AACTGT (coding strand)
Basic Elements of a Promoter Region in E. coli DNA:
- (-10)?
- 5’TATAAT3’/ 3’ATATTA5’
Basic Elements of a Promoter Region in E. coli DNA:
- what binds at -35 and -10?
sigma factor along with RNA poly
Basic Elements of a Promoter Region in E. coli DNA:
What is the one major sigma factor?
- sigma 70 –> it is the house keeping one generally most transcriptions created are done with this one
Basic Elements of a Promoter Region in E. coli DNA:
- heat shock genes?
- special sigma factors that are non 70…they promote the transcription of genes that will protect proteins from excess heat…protecting cells once they are in extreme environment
Sample Start and Stop sequences:
- What are two ways termination in prokaryotes can occur?
- transcription sequence folds creating a hairpin loop since it is complementary with itself which interferes with RNA sequence
- RHO factor: protein will come in bbd behind RNA poly and will basically push RNA poly out
Eukaryotic Poly II:
- what does GFTs stand for?
- general transcription factors
Eukaryotic Poly II:
-General transcription factors are required?
- TFIID with TBP
- TFIIA and TFIIB
- TFIIE, TFIIH and TFIIF with Poly II
Eukaryotic Poly II:
-RNA Polymerase II must be _____ so transcription can begin.
- phosphorylated
Eukaryotic Poly II:
What three things were studied to get a better understanding of the general transcription factors?
- Chicken ovalbumin
- Rabbit B-globin
- Mouse B-globin (major)
Eukaryotic Poly II:
Studied: Chicken ovalbumin, Rabbit B-globin and Mouse B-globin.
- Critical portion of promoter is where?
~24-32 bases upstream from initiation of transcription
Eukaryotic Poly II:
Studied: Chicken ovalbumin, Rabbit B-globin and Mouse B-globin.
- What are subunits of TFIID??
- TATA binding protein (TBP) and TATA- associated binging factors (TAFs)
Eukaryotic Poly II:
Studied: Chicken ovalbumin, Rabbit B-globin and Mouse B-globin.
- TATA binding proteins attach to? TAF’s binds to?
- TATA sequence
- promoter region
Eukaryotic Poly II:
Studied: Chicken ovalbumin, Rabbit B-globin and Mouse B-globin.
- TATA sequence is a what??
- promoter region that is hard to distinguish all the time (-10 region?)
Eukaryotic Poly II:
-General transcription factors determine what?
- the rate at which transcription is initiated at the promoter region
Eukaryotic Poly II:
- Steps of the general transcription factors?
- TFIID with TBP bind at TATAAbox(promoter element)
- TFIIA and TFIIB come in
- TFIIF with Poly II (together
- TFIIH: two functions…acts as a helicase and it phosphorylates RNA poly II
Eukaryotic Poly II:
- what is the complex called that contains the GFPs and RNA polymerase?
- Pre-initiation complex
* *TATA box= promoter region
Eukaryotic Poly II:
- CTD??
- carboxyl-terminal domain) from this complex permits modifications of pre-RNA strand that is coming out 7 aa’s in sequence
Eukaryotic Poly II:
- ELF?
- elongation factors permits RNA polymerase to proofread nt addition (so it can go back …stall …cut and go back again)
Primary Transcript Editing:
- Explain the 5’ capping!
L> what enzymes are involved
L>three functions of it?
- 5’ methylated guanosine cap
- -> phosphatase removes phosphates
- -> guanyl transferase–>GMP added
- methyl transferase–> methyl added
- Why does this need to happen???
1. these prevent the 5’ end from being digested by exonucleases which would cut it up.degrade it(acts as free end)
2. aids in transport out of nucleus(into cytoplasm via NPC)
3. plays role in trans+initiation of translation
- Why does this need to happen???
Primary Transcript Editing:
- what is the purpose of the 2 unsaturated regions ?
- dilute gene expression but are not part of the protein that it forms.
Primary Transcript Editing:
- Intron splicing?
- ~3500nt that need to be cut out_
- makes a huge mRNA strand smaller after splicing
Primary Transcript Editing:
-Poly A tail?
L> how is it added
L> function?
- 3’ end Poly A tail is added
- poly A polymerase adds the poly A tail
- adds ~250 A’s downstream from AAUAAA –> recognizes it and then tail is added
- f(x) prevents degradation in cytoplasm
Describe briefly the process of DNA–> mature mRNA
- Transcription by RNA poly II and caping make a primary transcript
- removal of 3’ end by nuclease and polyadenulation = processing intermediate
- endonucleolytic cleavage at splice junctions
- ligation of axons
- Mature mRNA
RNA Processing:
- what forms a spliceosome?
- snRNAs, mRNAs and protein
- heterogenous nuclear ribonucleoprotein
RNA Processing:
- what is the two step process?
- Lariat formation= loop
-5’ cleavage on intron - 3’ cleavage on intron
- 2 ends that introns that are removed are ligated together ( the two exons glued together after intron removal)
=====Mature RNA
RNA Processing:
- why do we want to remove introns?
- they intervene
- each exon has its own trait
- small nuclear RNA come in, in a specific order
RNA Processing:
- splicing of introns and fusing of axons is seen in eukaryotes and prokaryotes? Yes? No? Something different?
- ONLY seen in eukaryotes
RNA Processing:
- why do U1+U4 leave?
- U4 is an inhibitor
RNA Processing:
- Is U1 the first to come in?
- yes
- it comes in..binding to 5’ splice site.
RNA Processing:
- where does U2 come in at?
- branch point sequence
- U2 is the snRNA that oversees the lariat formation
RNA Processing:
- cluster of 3 snRNA?
- U5, U4 and U6
- U4 has an inhibiting role, prevents U6 from working (U6 cuts , cleaves)
- inhibit U6 until in right spot so you are leaving the right stuff (U4 does this).
- U5 +U6 allow axons to come together from working than U6 cleaves on 5’ splice site on intron.
- catalytic act that splices at 3’ end and there is now exon ligation and mature RNA forms. Intron gets chopped up and reused again.
mRNA Transport:
- transportation of an mRNA requires?
- the 5’ cap and 3’ poly A tail
mRNA Transport:
- transportation of an mRNA requires the 5’ cap and 3’ poly A tail…why?
- these are features that are recognized by receptor proteins located in NPC
- EIF eukaryotic protein initiating factors for translation. tRNA, ribosomal RNA’s all cme out
Genetic Code:
-RNA strand is in what direction?
5’-3’ –> read this way
Genetic Code:
- every three nt =?
- codon
- one amino acid
- a single aa is specified by a single codon
Genetic Code:
- codon is recognized by?
- tRNA’s anticodon
What are the requirements for Protein in synthesis in the cytoplasm?
- mRNA (template)
- a tRNA molecule with aa –> specifically coded for the codon it will attach to
- ribosome (small+large subunit)
- enzymes:
1. aminoacetly + RNA synthetase
2. peptidyl transferase - initiation factors and release factors (both euk + prok)
RNA coding:
- nucleotides found in?
- RNA
- Uracil, Adenine, Guanine and Cytosine
RNA coding:
- how many possible codons are there?
- 64
- four bases in 1, 4 bases in 2 , 4 names in 3
- There are 61 other codons and only 20 aa (code is degenerate meaning that one codon codes for more than one amino acid )
RNA coding:
- Three of the 64 possible codons are?
- Stop codons
- UAA, UAG and UGA
- *punctuated function meaning it marks the end
Reading Frame??Mutations?
- you need to write at a certain codon or else wrong codon.
- frameshift mutation if NT added that codes for a different protein.
tRNAs:
- act as?
- At least __ tRNA(s) per aa
- aa is linked to tRNA via?
- structure?
- adaptors
- one
- aminoacetly + RNA synthetase
- three hair pin loops w/ one aa accepting arm
- ACC (OH) on every accepting arm
The first two codon positions are the ?
- strictest (bc generally the last codon position doesn’t make a difference to the aa being coded
ex: CC– proline
The wobble base position is?
the third codon position
Ribosome:
- they are the catalyst of?
- two subunits?
- translation
- small subunit
- large subunit
Ribosome:
-describe the small + large subunit complex
-complex of RNA + protein
sm= bunds mRNA and tRNA
- large= catalyses peptide bond formation (bond between aa) by peptidyl transferase (ribosome not enzyme bc of its rRNA)
Ribosome:
- what are the three sites in the complex?
- E site
- P site
- A site
Initiation Process in Prokaryotes:
-Describe the process!
- IF(initiation factors) with small subunit will find the start codon on mRNA
- Shine delgarno sequence (sequence in mRNA, bind to some of the RNA in the small subunit to make sure smRSU is in the right place to put the first aa there)
- tRNA binds (IF1 and IF3 released)
- Large subunit binds (IF2 released)
Initiation Process in Prokaryotes:
-describe the function of IF1, IF2 and IF3
- IF1 facilitates attachment of smRSU to the RNA and helps tRNA initiator bind to site
- IF2 is necessary for allow initiator tRNA to bind (first tRNA that starts the translation every angle time –> carries methionine (AUG)
- IF3 prevents large subunit binding before it is suppose to
Initiation Process in Prokaryotes:
- what is the fmet equivalent in prokaryotes?
- IF3
Initiation Process in Prokaryotes:
- when do you have a tRNA in p site?
- when the tRNA is attached to a single aa
Initiation Process in Eukaryotes:
- how many complexes form?
- two complexes form independently
Initiation Process in Eukaryotes:
- what does mRNA bind to and how
-mRNA binds to 43S complex via EIF3 and EIF4G
Initiation Process in Eukaryotes:
-EIF4A prevents what
- hair pin loop
Protein Synthesis in Eukaryotes:
- explain the process
- initial tRNA (in P site) … tRNA enters at A site.
- tRNA in A site takes AA, peptide bond forms, translocation shifts one codon downstream , empty tRNA goes into E site, P site now has a tRNA with 2 AA.
- A is empty , tRNA enters A…taking the 2 aa’s and is now a tripeptide ..this process continues
Polyribosomes?
cluster of ribosomes, bound to a mRNA molecule progressing along the mRNA to synthesize the same protein
