Nucleic Acids Flashcards
What are nucleic acids composed of?
- Phosphate
- Pentose
- Purine or pyrimidine base
How are nucleic acids formed?
Two condensation reactions
What is the first condensation reaction of nucleic acids?
Phosphate is linked to carbon 5’ through a phospho-ester bond
What is the second condensation reaction of nucleic acids?
Purine or pyrimide (nucleic acid bases) is linked to carbon 1’ through a beta N-glycosidic bond
What is the difference between deoxyribose and ribose?
Deoxyribose: no O, just H at carbon 2’ (DNA)
Oxyribose: OH at carbon 2’ (RNA)
How many phosphate groups does a nucleotide contain?
One/nucleotide
Why are the numbers primed?
To distinguish them from the atoms in nitrogen bases
What are the 2 classes of nucleobases?
Derivatives of pyrimidine and purine
What are nucleobases chemically?
Nitrogen-containing heteroaromatic molecules
Name 2 characteristics of nucleobases.
- Planar or almost planar structures
- Absorb UV light around 250-270 nm
What # membered ring is purine? Pyrimidine?
Purine: 9 membered ring
Pyrimidine: 6
Name the 2 purines.
Adenine and Guanine
Name the 3 pyramidines.
Cytosine, Thymine, Uracil
Where is Thymine found? Where is Uracil found?
Thymine: DNA
Uracil: RNA
What does a nucleobase contain?
Nitrogenous base
What does a nucleoside contain?
Nitrogenous base and a pentose
What does a nucleotide contain?
Nitrogenous base, pentose and phosphate
How does polynucleotide polymerization occur?
3’ hydroxyl attacks the alpha-phosphate of another nucleotide (attached to carbon 5’) and releases a pyrophosphate
What kind of linkage do you form between 2 nucleotides? Between what?
Phosphodiester linkage
- 3’ and 5’
What is the convention for writing DNA/RNA polymers?
- Top to bottom, left to write
- 5’ end to 3’ end
What is the backbone of DNA/RNA polymer made of?
Alternating phosphate and sugar groups
Which end of DNA/RNA polymers are phosphorylated? Which aren’t?
Phosphorylated: 5’ end
NOT phosphorylated: 3’ end
What are oligonucleotides? Polynucleotides?
Oligonucleotides: less than 50 nucleotides
Polynucleotides: > 50
What are DNA/RNA polymers sensitive to? Which is more sensitive? Why?
- Sensitive to alkaline (base) hydrolysis
- RNA is more sensitive due to 2’ OH
What is the shorthand representation of phosphodiester linkage?
3’-5’
In the early 1900s, scientists believed that proteins were responsible for the storage and transport of nucleic acids. Name the 3 discoveries that proved them wrong.
- Griffit: I don’t know if proteins are responsible created a doubt, maybe it is nucleic acids
- Avery: proved it was DNA, could not rule out proteins
- Hershey: ruled out proteins completely
What was Griffith? What year did he conduct his experiment in? What was his discovery? Why is it only doubt?
- Bacteriologist
- 1928
- Transforming Principle
- Some proteins are heat resistant, only creates doubt
What is the transforming principle by Griffith?
transforming material (unlikely proteins)
crossed from the heat killed virulent S strain to the
non-virulent R live strain
- Mixture of heat-killed virulent and live nonvirulent bacteria
- dead bacteria of a virulent type and live bacteria of a non-virulent type were both injected in mice –> dead mice + virulent bacteria could be extracted from the dead mouse
What did Avery do? What was his conclusion?
Mixing of purified fractions from virulent with live non-virulent bacteria
and isolation of S-virulent strain on petri dish
- DNase-treated but NOT protease-treated
- Transforming factor is DNA
What was Hershey’s conclusion? How is protein ruled out?
Injected material from the Bacteriophage was P32 labeled and nucleic acids are the ones that contain phosphorus! DNA is responsible for the storage and transfer of genetic material
- Amino acids (cysteine and methionine) both have S groups, but radioactive S were not transferred, so protein is not responsible for the storage and transfer of genetic material
What was Crick and what was Watson? What did they want to discover? What did they discover?
Crick: physicist
Watson: molecular biologist
- Wanted to determine the structure of nucleic acids
- Evidence that nucleic acids can store genetic material in the sequence of their bases
What information helped Crick and Watson in their discoveries? (3)
1) Nucleic acids are helical in structure (2 helices) - Rosalind Franklin
2) The two strands are separated by 3 A (Rosalind Franklin)
3) Chargaff’s Rule: A+C = T+G, nucleotides have a constant ratio
What did Crick and Watson state in their publication that explained the secret of life?
Not only storing, but possible copying mechanism for the genetic material
How many hydrogen bonds do AT have? How many hydrogen bonds do GC?
AT: 2 hydrogen bonds
GC: 3 hydrogen bonds
Do you need more energy to break GC or AT bonds?
GC, since 3 hydrogen bonds
What is the distance between nucleobases?
3 A
Is the double helix right or left handed?
Right
What grooves does the helix contain? What is the pitch from one groove to the next? What is the width of the double helix? How many nucleotides per helix turn?
- Major and minor
- 36 A
- 20 A
- 10.4 nucleotides/helix turn
Where do the phosphates and sugar groups point? Where do the nucleic acids point?
Phosphate and sugar: outside
Nucleic acid: inside
How are the bases held together? (2)
- Hydrogen bonds
- Van der Waals interaction
The bases are separated to each other along the helix axis by what?
3.4 A
Describe the strands of the helix.
- 2 anti-parallel
- complimentary (5’—3’ & 3’—5’)
How does DNA get converted to protein?
- Replication of DNA
- Transcription -> RNA
- RNA -> Protein through translation
Name the 3 postulated methods of DNA replication.
1) Semi-Conservative
2) Conservative
3) Dispersive
What is the major disadvantage of conservative DNA replication?
No genetic variation, not a good evolutionary concept
What would happen if DNA replication were to occur by a conservative mechanism?
two original template strands would stay together, and would instead produce an entirely new double-stranded copy.
Is DNA replication unidirectional or bidirectional? Explain how we found this out.
- Bidirectional
- Take bacteria and grow in small amount of THYMIDINE (since DNA), spread on slide, put it on a film, the more you expose it, the darker the signal once you develop the film
- Dark lines on both sides = bidirectional
Name the 4 components needed for DNA replication.
1) DNA template
2) Primer strand
3) dNTPs (deoxyribonucleoside triphosphate)
4) Various bacterial fractions
What is the importance of the primer strand?
Provides you with a 3’ hydroxyl end, the end that does the nucleophilic attack on dNTPs, which is required for DNA replication
What do DNA polymerases catalyze? In what direction?
The stepwise addition of deoxyribonucleotides (5’-3’)
What characteristics of DNA polymerase I and DNA polymerase III share?
1) It has 5’ — 3’ polymerase activity
2) It has 3’ — 5’ exonuclease activity (proof reading)
3) Requires a DNA template
What is an exonuclease?
Removes nucleotides from the end of a polynucleotide molecule
What does DNA polymerase I have that DNA polymerase III doesn’t? What does it do?
- 5’—3’ exonuclease activity (removes added primers)
How do DNA polymerase I and DNA polymerase III compare in terms of processing?
1: moderately processive (dissociates after adding 20 dNTPs)
3: synthesizes 1000 dNTPs/second
The three activities of DNA polymerase I occur where? How many subunits make up DNA polymerase III?
- On a single polypeptide (I)
- 10 subunits (III)
What does Helicase do? Does it require energy?
Unwinding of the DNA double Helix
Requires energy
What do single strand binding proteins do?
To keep DNA strands separate, to not form back the helix
What does topoisomerase do?
Releases the stress of DNA unwinding through transient DNA cut and ligation
What does primase not require to add complimentary base pairs? It is a must for what? What is it made of? What is the extension of DNA done by?
- Does not require free 3’OH group to add complimentary base pairs
- Must for Leading and Lagging Strand
- RNA primer, extension of DNA done by DNA polymerase
How many primers do you need for the leading strand?
One
The lagging strand synthesis is unique. How does it occur? What are these called?
- Occurs in a fragmented fraction
- Okazaki fragments
How many primers do you need for the lagging strand? How does DNA synthesis occur?
- You need a primer for EVERY fraction
- Then DNA copies, then primer, then DNA
What removes the RNA primer? What does it put instead?
5’-3’ exonuclease activity of DNA polymerase I
- Fills the gap with DNA
What joins the DNA fragments on the lagging strand?
DNA ligase
What enzymes act as the rate limiting step for DNA replication?
Helicase and primase
