DNA Repair
How are mistakes made during DNA replication fixed?
DNA polymerase can back up and fix the bases it wrongly added, otherwise a mismatch repair enzyme can fix it (MMR). A mistake is an improper base pair, recognized because the DNA helix has become distorted.
What is a promoter and how does it affect DNA transcription?
It is a region of DNA before the start of the gene that binds transcription factors (in eukaryotes) that recruit RNA polymerase (what makes the RNA from the DNA template)
What terminates transcription?
At the termination sequence, RNA dissociates from DNA template
(due to formation of a self-base pairing loop that pulls RNA polymerase “off its tracks” (in prokaryotes)
Stem loops forms a stable structure that slows the RNA polymerase down, stretch of weak AU bps makes it easy for mRNA to tear away)
What are the three types of RNAs used in translation and what role do they each play?
a. mRNA- product of transcription that specifies sequence of aa in the polypeptide
b. rRNA- major component or ribosomes (also the product of transcription)
c. tRNA- links aa to codons in the mRNA (also the product of transcription)
What is translation? What makes it happen and where does it happen?
The process by which mRNA information is used to create a protein via ribosomes located in the cytosol.
How does UV radiation damage DNA?
Dimerizes adjacent thymines (creates covalent bond), causing bulge in DNA strand that prevents replication and transcription from occurring
What happens during transcription? Where does it occur and why might it occur?
mRNA is synthesized using DNA as a template using RNA polymerase, occurs at genes as necessary based on needs/type of cell
What happens to the RNA after it has been transcribed in eukaryotes? In prokaryotes?
Eukaryotes- RNA is processed, shipped out of nucleus, then translated
Prokaryotes- RNA is immediately translated as it is transcribed
What are codons and how does the tRNA use them?
How is translation initiated?
Small subunit binds (towards 5’ end, Shine-Dalgarno sequence in bacteria and 5’ cap in eukaryotes), methionine/AUG tRNA binds, large subunit binds
Aided by initiation factors
Is the entire mRNA produced by transcription translated?
No, only the coding region (not the UTR)
What are transcription factors? Why might it be useful that the RNA polymerase can’t bind to the promoter without them?
Proteins that bind near the promoter (TATA box) that allow/help RNA polymerase to bind
Allow for regulation. For example, some hormones bind to intracellular receptors and enter the nucleus so that they can bind to the promoter. This allows body to send signals about what genes need to be transcribed. (only applicable in euks)
What is a frameshift mutation and how does it affect translation?
Insertion or deletion of a small number of bases that throws off the reading frame, causing all sequential amino acids to be wrong
Describe what happens in each site of the ribosome.
A: holds upcoming tRNA before its attached amino acid is joined to the polypeptide chain and binds release factor that ends translation and dissociates ribosome
P: tRNA’s polypeptide chain is joined to the aa in the A site
E: hold tRNA with no amino acid (uncharged) before it exits the ribosome
What proteins are necessary for the initiation of transcription?
RNA polymerase, transcription factors or sigma factor
How does nucleotide excision repair work?
a. Enzymes detect an irregularity in DNA structure and cut the damaged strand.
b. An enzyme excises nucleotides on the damaged strand.
c. DNA polymerase fills in the gap in the 5’ - 3’ direction using other strand as template
d. DNA ligase links the new and old nucleotide
What is a sigma factor?
Protein that helps RNA polymerase begin transcription then dissociates (it helps RNA pol bind to a specific promoter sequence; there are diff sigma factors that lead to transcription of diff genes), found only in prokaryotes.
How is RNA processed and why?
Addition of 5’ cap and 3’ polyA tail, in order to increase stability of RNA (poly-A tail) and promote its translation (5’ cap)
RNA splicing- remove introns and splice exons together via enzyme spliceosome (made up of protein and small nuclear ribonucleoproteins aka snurps)
How are tRNAs “charged?” How is specificity ensured in this process?
What are the three types of single base (point) mutations? How does each affect translation of the gene?
Point mutation is a single nucleotide base pair change
●Synonymous (silent) mutations; no change in the amino acid
●Nonsynonymous (missense) mutations; change in the amino acid
●Nonsense mutations; where the change results in a stop codon
Determine which polymerase the researcher has isolated based on the given information
a. Protein A
i. Synthesizes a nucleic acid from a single strand of DNA
ii. Functions when helicase activity is suppressed
b. Protein B
i. Can correct errors
ii. Uses dNTPs
c. Protein C
i. Synthesizes the following nucleic acid from a primer: ACGGGCAG
ii. Nucleotides used lack a 2’ OH
d. Protein D
i. Associates with a sigma factor
ii. Does not use thymine
e. Protein E
i. Uses an RNA template
ii. Adds nucleotides to the lagging strand template
f. Protein F
i. Found near the TATA box
ii. Adds nucleotides to the 3’ end of a growing strand
g. Protein G
i. Never found in the nucleus
ii. May transcribe multiple genes at once
iii. No lagging strand is formed
Determine which polymerase the researcher has isolated based on the given information
a. Protein A RNA pol (I-III, multiple RNA pol can do this)
i. Synthesizes a nucleic acid from a single strand of DNA
ii. Functions when helicase activity is suppressed
b. Protein B DNA pol (III)
i. Can correct errors
ii. Uses dNTPs
c. Protein C DNA pol (I-V, multiple DNA pol can do this)
i. Synthesizes the following nucleic acid from a primer: ACGGGCAG
ii. Nucleotides used lack a 2’ OH
d. Protein D RNA pol, prokaryotic
i. Associates with a sigma factor
ii. Does not use thymine
e. Protein E telomerase
i. Uses an RNA template
ii. Adds nucleotides to the lagging strand template
f. Protein F RNA pol (II), eukaryotic
i. Found near the TATA box
ii. Adds nucleotides to the 3’ end of a growing strand
g. Protein G RNA pol, prokaryotic
i. Never found in the nucleus
ii. May transcribe multiple genes at once
iii. No lagging strand is formed
What is alternative splicing? Why might it occur?
Different proteins can be made from a single gene, depending upon which introns are removed and which exons are present when the RNA transcript is spliced together.
What does it mean to be polycistronic?
An entire operon (composed of multiple genes) is transcribed as a single mRNA. Then ribosomes begin translation at the Shine-Dalgarno sequence at the beginning of each gene and translates until the stop codon of each gene to produce the different polypeptides to fold into functional proteins.
In what direction is mRNA read by the ribosome?
5’ --> 3’
***this is an important difference between DNA replication/transcription reading direction and translation***