Exam 4 Nucleotides and Nucleic Acids

Question 1

What are nucleic acids and their functions?

Answer 1

Nucleic acids are polymers of nucleotides. They are used for storing and transmitting genetic info (DNA, mRNA), processing genetic info (ribozymes), and protein synthesis (tRNA, rRNA)

Question 2

What are nucleotides used in the monomer form for?

Answer 2

It is used for energy for metabolism (ATP, enzyme cofactors (NAD+), and signal transduction (cAMP)

Question 3

What is the difference between nucleotide and nucleoside?

Answer 3

• Nucleotide: nitrogenous base, pentose and phosphate (attached to pentose 5’C, impt for backbone)
• Nucleoside: nitrogenous base and pentose

Question 4

What is the cyclic format of the carbon and nitrogen atoms of pyrimidine and purine?

Answer 4

Pyrimidine: 1’ starts at center N and goes to the left
Purine: 1’ starts at left N and goes to the right

Question 5

Phosphate group consists of?

Answer 5

• negative charge at neutral pH
• attached to 5’ position (nucleic acids built using 5” triphosphates of nucleotide - ATP, GTP, TTP, CTP)
• 2 of 3 phosphates used to build nucleic acid form a leaving group
• completed nucleic acid has one phosphate moiety per nucleotide

Question 6

How do pentose forms differ between RNA and DNA?

Answer 6

• RNA: b-D-ribofuranose
• DNA: b-2’-deoxy-D-ribofuranose
  Different puckered conformation of sugar ring possible -> form is due to sugar and phosphate group

Question 7

What are nitrogenous bases?

Answer 7

• derivatives of pyrimidine or purine
• nitrogen-containing heteroaromatic
• planar/almost planar
• absorb UV light at 250-270nm
• neutral at pH 7, good H-bond donors/acceptors
• In DNA: C, A, G, T
• In RNA: C, A, G, U

Question 8

What is the b-N-Glycosidic Bond?

Answer 8

Pentose ring attached to the nitrogenous base via N-glycosidic bond (bond is formed to anomeric carbon of sugar in b-configuration)
- In pyrimidines: to position N1
- In purines: to position N9

Question 9

What are polynucleotides?

Answer 9

Covalent bonds formed via phosphodiester linkages (neg. charged backbones)
- DNA backbone: stable, hydrolysis accelerated by DNAse
- RNA backbone: unstable (in water: RNA lasts few years, in cells: mRNA lasts few hours)
- Linear: no branching/cross-links
- Directionality: 5’ - 3’ (5’ end is diff from 3’ end)

Question 9

MInor nucleosides in RNA

Answer 9

Inosine: found in the wobble position of anticodon in tRNA (provides richer genetic code)
- made by: de-animating adenosine
Pseudouridine: in tRNA and rRNA
- in eukaryotes/eubacteria
- made from uridine by enzymatic isomerization (after RNA synthesis)
- stabilize structure of tRNA

Question 9

When is the b-N-glycosidic bond stable?

Answer 9

Stable towards hydrolysis especially in pyrimidines

Question 9

How does bond cleavage of b-N-glycosidic bond happen?

Answer 9

catalyzed by acid

Question 9

What limits the bond rotation possibilities?

Answer 9

Ring pucker limits the angle of torsion, c, possible for the N-glycosidic bond between the nitrogenous base and pentose

Question 9

Conformation around N-glycosidic bond

Answer 9

Free rotation around bond in free nucleotides
- syn conformation: angle near 0 (leans to right) -
- anti conformation: angle near 180 (leans to left) - found in normal B-DNA & purines can only be anti

Question 9

Hydrolysis of RNA

Answer 9

• RNA unstable under alkaline conditions
• Hydrolysis catalyzed by enzymes (RNAse)

Question 9

Why can nucleobases absorb UV light at 250-270nm?

Answer 9

It is due to n -> n* electronic transitions

Question 10

Minor nucleosides in DNA

Answer 10

Modification done after DNA synthesis
- 5-Methylcytosine: in eukaryotes and bacteria
- N6-methyladenosine: in bacteria but not eukaryotes
- Epigenetic marker: to mark own DNA to degrade foreign DNA (prokaryotes), mark which genes should be active (eukaryotes)

Question 10

What are the Hydrogen Bonding Interactions of bases?

Answer 10

Two bases can H-bond to form a base pair (large # possible for monomers)
- Watson-Crick base pairs predominate in double-strand DNA ( A &T, C&G, Purine & Pyrimidine)

Question 10

RNAse enzymes?

Answer 10

• S-RNase in plants: prevents inbreeding
• RNAse P: ribozyme that processes tRNA precursors
• Dimer: enzyme that cleaves double-stranded RNA into oligonucleotides

Question 10

What is the complementarity of DNA strands?

Answer 10

• Two chains sequences differ
• They are complementary
• They run antiparallel

Question 10

How does replication of genetic code happen?

Answer 10

1) Strand separation
2) Each strand serves as template for synthesis of new strand
3) Synthesis catalyzed by DNA polymerases (enzymes)
4) Newly made DNA molecule has one daughter and one parent strand

Question 10

What is mRNA?

Answer 10

Code carrier for the sequence of proteins (transfers genetic info from DNA with tRNA)
- Made from DNA template
- Single strand
- Contains Ribose (not deoxyribose) and Uracil
- one mRNA can code for more than one protein

Question 10

What are palindromic sequences and their function?

Answer 10

nucleic acid sequence in DNA or RNA that reads the same in both directions on complementary strands (opposite strands)

Question 11

How are complex structures stabilized?

Answer 11

Complex structures are stabilized by non-watson-crick base-pair interactions

Question 12

What happens in DNA Denaturation?

Answer 12

• Covalent bonds stay intact (genetic code intact)
• Hydrogen bonds are broken (strands separate)
• Base stacking is lost, which increases UV absorbance

Question 13

What induces denaturation and how can it be reversed?

Answer 13

• Induced by: high temp or change in pH
• Reversible by: annealing
  (This is the basis for PCR)

Question 14

What happens in thermal DNA denaturation (melting)?

Answer 14

At elevated temps: the two DNA strands dissociate (not a double helix anymore)
- At lower temps: the two strands re-anneal
- UV absorbance increases as the molecule falls apart (usually monitored at 260nm)

Question 15

What are the factors that affect DNA denaturation?

Answer 15

• Midpoint of melting (Tm) depends on base composition: High GV = increased Tm
• Tm depends on DNA length: longer DNA = higher Tm
• Tm depends on pH and ionic strength: high salt = increased Tm

Question 16

Do AT-rich regions of DNA melt at a different temperature than GC-rich regions?

Answer 16

AT-rich regions melt at a lower temperature than GC-rich regions

Question 17

Molecular Mechanisms of Spontaneous Mutagenesis: Deamination and Depurination

Answer 17

Deamination:
- very slow reactions, large # of residues, significant net effect
Depurination:
- N-glycosidic bond is hydrolyzed
- significant for purines (10,000 lost/day)

Question 18

Molecular mechanisms of oxidative and chemical mutagenesis

Answer 18

Oxidative damage:
- hydroxylation of guanine
- mitochondrial DNA most susceptible
Chemical alkylation:
- methylation of guanine

Question 19

Molecular mechanisms of radiation-induced mutagenesis

Answer 19

• UV light: induces dimerization of pyrimidines - main mechanism for skin cancers
• Ionizing radiations (xrays/yrays): causes ring opening and strand breaking

Question 20

What are accumulations of mutations linked to?

Answer 20

Cells can repair some modifications but others cause mutations
- It links to aging and carcinogenesis

Question 21

What are A, B, Z DNA?

Answer 21

• A-DNA is a right-handed double helix made up of deoxyribonucleotides. …
• B-form DNA. B-form DNA is a right-handed double helix (discovered by Watson and Crick based on the X-ray diffraction patterns)
• Z-form DNA. Z-form DNA is a left-handed double helix.

Question 22

What are Chargaff’s rules?

Answer 22

of adenine = # of thymine
# of cytosine = # of guanine
# of purine = # of pyrimidine
(1:1 ratio of bases)