DNA Structure & Base Pairing
What is the difference between a nucleoside and a nucleotide?
A nucleoside is a base + sugar; a nucleotide is a nucleoside with one or more phosphate groups.
What does it mean for DNA strands to be antiparallel?
One strand runs 5′→3′ and the other 3′→5′, enabling base pairing.
Which DNA structures support synthesis and why?
(a) Closed ss circle
(b) Closed ds circle
(c) ss paired with linear strand with 3′-OH
(d) ds linear with free 3′-OH
Only (c): ssDNA paired to a linear strand with 3′-OH allows synthesis.
How does alternative splicing increase genomic capacity?
It allows one gene to produce many mRNAs and proteins.
What are key features of the genetic code?
The code is triplet, non-overlapping, universal, degenerate, and has no punctuation.
This DNA-processing enzyme uses dNTPs as activated intermediates and releases pyrophosphate.
DNA polymerase
What is a Watson-Crick base pair?
A–T and G–C base pairs held together by hydrogen bonds.
Why is the double helix stable despite weak bonds?
The large number of weak hydrogen bonds and base stacking together stabilize the helix.
What defines a retrovirus?
A virus with RNA as its genome that reverse-transcribes into DNA.
What is gene expression?
The process of converting a gene into RNA and/or protein.
Given the DNA coding strand 5′-ATCGTACCGTTA-3′, what is the corresponding mRNA sequence transcribed during transcription?
The mRNA sequence is identical to the DNA coding strand except that thymine (T) is replaced with uracil (U) in RNA.
5′-AUCGUACCGUUA-3′
This enzyme uses AMP–DNA intermediates to seal DNA breaks.
DNA ligase
If DNA contains 20% thymine, what are the percentages of the other bases?
A = 20%, G = 30%, C = 30%.
What are the 3 DNA helical forms and one difference between them?
A-DNA (short, wide), B-DNA (standard), Z-DNA (left-handed).
What is the role of ribonuclease in reverse transcription?
It removes the RNA strand from RNA–DNA hybrids so DNA can be synthesized.
What is a consensus sequence?
A sequence representing the most frequent bases at each position in aligned sequences; helps identify key regulatory regions.
Why is RNA more easily hydrolyzed than DNA under alkaline conditions?
RNA has a reactive 2′-OH that attacks (nucleophile) the phosphodiester backbone under basic conditions.
This topological enzyme uses a covalent tyrosine–DNA intermediate to temporarily break DNA strands and rejoin strands.
Topoisomerase
Write the complementary 5′→3′ sequences for:
(a) GATCAA
(b) TCGAAC
(c) ACGCGT
(d) TACCAT
(a) TTGATC
(b) GTTCGA
(c) ACGCGT
(d) ATGGTA
Define template and primer in DNA synthesis.
Template: guides the new strand. Primer: provides the 3′-OH to start synthesis.
Compare DNA and RNA polymerase in:
(a) Precursors
(b) Direction
(c) Template conservation
(d) Primer requirement
(a) DNA: dNTPs, RNA: NTPs;
(b) Both 5′→3′;
(c) Both preserve template;
(d) DNA needs primer, RNA does not.
What happens if sigma factor can bind the −10 region alone?
It would block RNA polymerase holoenzyme binding and inhibit proper transcription.
Why is RNA more prone to nucleophilic attack than DNA?
The 2′-OH allows cyclic intermediates (compounds) to form in RNA but not in DNA.
This enzyme must be reactivated in cancer cells to avoid telomere shortening.
Telomerase
Why are GC and AT the only base pairs allowed in the double helix?
Two purines are too bulky, and two pyrimidines are too small. GC and AT pairings maintain helix geometry.
Why does DNA replication halt without topoisomerase?
Overwinding halts replication unless topoisomerase relieves the tension.
What’s the difference between coding and template DNA strands?
Coding strand matches mRNA (T/U difference); template strand is used to make RNA.
What happens if sigma can’t dissociate from core polymerase?
Transcription would slow down, and the enzyme would struggle to bind alternative σ factors.
How does cordycepin inhibit RNA synthesis?
It lacks a 3′-OH, blocking 3′→5′ phosphodiester bond formation, so once incorporated, RNA synthesis terminates.
In telomerase-deficient cells, chromosomes undergo this fate.
Chromosomes undergo end-to-end fusions due to exposed DNA ends.
Why does heat melt DNA?
Heat disrupts hydrogen bonding and base stacking that stabilize the double helix.
Which sugar position excludes ribonucleotides during DNA synthesis?
DNA polymerase contacts the 2′-OH to exclude ribonucleotides.
How do mRNA, tRNA, and rRNA differ in structure and function?
mRNA: carries genetic info
tRNA: brings amino acids
rRNA: forms ribosomes
Where does transcription likely begin in this DNA sequence?
5′-TATAATCGGCGATGCATGCTAGGATCCCGCTT-3′
The A immediately after the TATA sequence (the TATA box).
A in "ATCGGCG..."
What is degeneracy in the genetic code?
Most amino acids are encoded by more than one codon.
Guanine oxidation can lead to this phenomenon in triplet repeat regions.
Strand slippage and repeat expansion
What properties of DNA make it ideal for genetic material?
DNA is stable, mutable, replicable, and reliably stores large amounts of information.
What direction does DNA polymerase synthesize, and why?
DNA synthesis proceeds 5′→3′ so that the 3′-OH can attack the α-phosphate of incoming dNTPs.
Why is RNA synthesis not monitored as strictly as DNA synthesis?
RNA errors affect only one transcript; DNA errors are permanent and heritable.
Why does heparin inhibit transcription?
It is highly anionic and mimics DNA’s negative charge, allowing it to bind RNA polymerase’s lysine and arginine residues.
What is the benefit of a degenerate code?
It reduces the impact of point mutations on protein function.
Why are there multiple DNA repair pathways?
The variety of damage types requiring specialized mechanisms.
What is positive supercoiling and how does it stabilize DNA?
Positive supercoiling stabilizes DNA at high temperatures by overwinding the helix.
Why is AMP, CTP, GTP, and UTP needed for DNA replication?
These NTPs are needed to synthesize RNA primers via primase.
Why is DNA synthesis faster than RNA synthesis?
Only a fraction of the genome is transcribed at a time, so speed is not essential for RNA synthesis.
Why does abortive cycling occur during transcription initiation?
Initial RNA-DNA hybrids are unstable and can't prevent early strand loss, leading to frequent early termination.
What is exon shuffling and why is it evolutionarily useful?
Rearranging exons creates new proteins and speeds up evolution.
What is the function of AMP-PNP in topoisomerase II analysis?
AMP-PNP prevents DNA release, showing ATP hydrolysis is essential for function, since ATP hydrolysis is needed for DNA release.
Why does DNA need Mg²⁺ ions?
Mg²⁺ shields negative charges on phosphate groups to reduce repulsion.
If Tw = 48 and Wr = 3, what is Lk? If Tw = 50 and Lk = 51, what is Wr?
Lk = Tw + Wr; so: Lk = 48 + 3 = 51; Wr = Lk - Tw = 1.
What does the similarity between polymerase active sites suggest?
Convergent evolution: similar function shaped similar catalytic sites.
What happens if topoisomerase II transcription is self-regulated?
If topoisomerase II transcription is self-regulated, too much negative supercoiling inhibits its own transcription by distorting the promoter, acting as a built-in mechanism to keep DNA topology balanced.
What increases proteome complexity beyond the genome?
Alternative splicing, RNA editing, and post-translational modifications.
Match components to function:
Functions:
(a) Replication
(b) Transcription
(c) Translation
Components:
RNA pol
DNA pol
Ribosome
dNTP
tRNA
NTP
mRNA
Primer
rRNA
RNA pol → (b)
DNA pol → (a)
Ribosome → (c)
dNTP → (a)
tRNA → (c)
NTP → (b)
mRNA → (b), (c)
Primer → (a)
rRNA → (c)
Why does DNA synthesis occur in the 5′→3′ direction?
Because the 3′-OH attacks the α-phosphate of the incoming nucleotide.
What explains DNA spinning speed at the replication fork in E. coli?
E. coli synthesis rate ≈ 0.34 μm/s; DNA rotation ≈ 96.2 revolutions/sec at the fork.
Why are proofreading products of RNA polymerase mostly dinucleotides?
RNA polymerase often backtracks before cleavage producing dinucleotides, not single nucleotides.
What explains the effect of negative supercoiling on transcription?
Supercoiling may alter −10 and −35 element alignment, acting as self-regulation to prevent overexpression. It prevents overexpression of topoisomerase II by distorting its own promoter region.
How can mRNA be separated from other RNAs in eukaryotic cells?
By using oligo(dT) or oligo(U) affinity columns to bind mRNA poly(A) tails.
Match the components in the right-hand column with the appropriate process in the left-hand column.
Processes:
(a) fMet
(b) Shine–Dalgarno
(c) Introns
(d) Exons
(e) pre-mRNA
(f) mRNA
(g) Spliceosome
Components:
1. Joins exons
2. removed
3. first amino acid
4. joined
5. final message
6.start site
7. unprocessed message
fMet → first amino acid
Shine-Dalgarno → start site
Introns → removed
Exons → joined
pre-mRNA → unprocessed message
mRNA → final message
Spliceosome → joins exons