2. Nucleic Acids

Question 1

what are the types of nucleic acids, what are they composed of

Answer 1

Types of nucleic acids: DNA, RNA

composed of nucleotides:
- contains a base - Adenine, cytosine, guanine, thymine, uracil
- a sugar - deoxyribose and ribose
- a phosphate

the structure of nucleotides incorporated into the polymer determines the structure of the nucleic acid

Question 2

DNA

Answer 2

A polymer of deoxynucleotides whose sequence of bases encodes genetic information in all living cells

Question 3

RNA

Answer 3

A polymer of ribonucleotides, such as messenger RNA (mRNA), transfer RNA (tRNA), and ribosomal RNA (rRNA)

Question 4

nucleotide

Answer 4

• compound consisting of a nucleoside esterified to one or more phosphate groups
• monomeric units of nucleic acids

Question 5

nucleoside

Answer 5

compound consisting of a nitrogenous base linked to a 5-carbon sugar (ribose or deoxyribose)

Question 6

deoxynucleotide

Answer 6

nucleotide in which the pentose is 2’-deoxyribose

Question 7

2 common sugars

Answer 7

deoxyribose and ribose

Question 8

bp and kb

Answer 8

bp - base pairs
kb - kilo base pairs (1000 base pairs)

Question 9

oligonucleotide

Answer 9

A polynucleotide consisting of a few nucleotide residues

Question 10

B-DNA

Answer 10

The standard conformation of double-helical DNA

Question 11

stacking interactions

Answer 11

The stabilizing van der Waals interactions between successive (stacked) bases in a polynucleotide

Question 12

melting temperature (Tm)

Answer 12

The midpoint temperature of the melting curve for the thermal denaturation of a macromolecule.
For a lipid, the temperature of transition from an ordered crystalline state to a more fluid state

Question 13

denaturation and renaturation

Answer 13

denaturation: the loss of ordered structure in a polymer
i.e disruption of native conformation (unfolded polypeptide), unstacking of bases, separation of strands in a nucleic acid

renaturation: the refolding of a denatured macromolecule so as to regain its native conformation

Question 14

anneal

Answer 14

to allow base pairing between complementary single polynucleotide strands so that double-stranded segments form

Question 15

Distinguish between properties of purine and pyrimidine bases.

Answer 15

Purines: Adenine (A), Guanine (G)

• double ring structure, basically planar, slight pucker in purine base
• poorly soluble in water
• largely hydrophobic with some polar groups (ability to form H-bonds)

Pyrimidines: Cytosine (C), Thymine (T), Uracil (U)

• heterocyclic
• aromatic, electron delocalization

Question 16

What are the 5 common bases? Which are classified as pyrimidines and which are purines?

Answer 16

Purines: Adenine (A), Guanine (G)

Pyrimidines: Cytosine (C), Thymine (T), Uracil (U)

T - DNA
U - RNA

Question 17

Is the sugar phosphate backbone polar or non polar?

Answer 17

polar

Question 18

Name and distinguish between the two sugars that are incorporated into nucleotides

Answer 18

Ribose: found in ribose, has a -OH attatched to the 2’ carbon of the sugar ring in replace for a H present in deoxyribose

Deoxyribose: found in DNA, lacks -OH on the 2’ carbon (deoxy)

• both are furanose rings (5 membered with an oxygen)

Question 19

naming of nucleosides and nucleotides

Answer 19

Nucleosides:

• -osine in purines replaces -ines
  (eg. adenine + ribose = adenosine)
• pyrimidines → end in -idine
  (eg. cytosine + ribose = cytidine)
• deoxy- is added as a prefix for nucleosides with deoxyribose sugar
  (eg. thymine + deoxyribose = deoxythymidine

Nucleotides:
* nucleoside name + “5’ ____ - phosphate”
* ____ is the prefix for amount of phosphate’s in the phosphate group (mono-/di-/tri-/tetra-)
* phosphates attatched to multiple carbon/OH groups are described using bis-/tris- prefixes along with including the atom number that the phosphate is attatched to
(eg. adenosine 2’, 5’ bisphosphate)

5’ refers to carbon in the sugar ring, most of the time attatched to 5’ carbon but can be C2 or C3 as well

Question 20

compare the structure of nucleosides with the structure of nucleotides

Answer 20

• nucleosides: sugar + base, no phosphodiester bond
• nucleotide: sugar + base + phosphate group, contains phosphodiester bond

Question 21

Define the term nucleic acid

Answer 21

• also known as a polynucleotide
• polymer of nucleic acids
• major nucleic acids: deoxyribonucleic acid (DNA), ribonucleic acid (RNA)

Question 22

explain why nucleic acids are said to have a “sense of direction”

Answer 22

• phosphodiester bonds in nucleic acids create asymmetric structures with a “sense of direction” →
• phosphodiester bonds link the 3’ carbon of one sugar and the 5’ carbon of the next sugar → creates a repeating sugar-phosphate backbone.

5’ end → 3’ end

Question 23

What are the 5’ and 3’ ends of nucleic acid base sequences?

Answer 23

• The 5’ end of the nucleic acid chain has a free phosphate group attached to the 5’ carbon of the sugar.
• The 3’ end has a free hydroxyl (-OH) group attached to the 3’ carbon of the sugar
• nucleotides joined by phosphodiesters are described as residues

Question 24

What is the primary (1°) structure for a nucleic acid?

Answer 24

the sequence of nucleotide residues; the order of monomers in a polymer

Question 25

abbreviations for **primary structures** of nucleic acids

Answer 25

based on the first letter of each nucleotide written in sequence from 5' → 3' by convention e.g: 5' - ATGCAATG - 3' can also be written as: ATGCAATG, 5'-dAdTdGdCdCdCdTdG-3', dAdTdGdCdCdCdTdG (DNA) * T indicates the nucleic acid is likely DNA * U indicated the nucleic acid is likely RNA

Question 26

phosphoesters, phosphodiesters, phosphoanhydrides

Answer 26

1. phosphoesters: (phosphate esters), link carbons to phosphate groups 2. phosphodiesters: link two different carbons to one phosphate group 3. phosphoanhydrides: link two phosphates to each other (within a phosphate group)

Question 27

describe and compare the primary structure of RNA and DNA

Answer 27

Both: have a net (-) charge, a polar backbone, a phosphodiester linkage following from the 5' end of the nucleotide to the 3' end, and a free 3' -OH group at the 3' end RNA: contains uracil instead of thymine and is a little more polar due to the extra -OH on the 2' carbon of the nucleotide sugars (ribose) DNA: contains thymine instead of uracil, slightly less polar than RNA due to H+ on 2' carbon

Question 28

explain why RNA is susceptible to alkaline hydrolysis whereas DNA is resistant

Answer 28

phosphodiester bond spontaneously hydrolyze (phosphodiesterase) at pH>10 In DNA, the absence of a 2' OH prevents the reaction. This makes DNA more stable than RBA under alkaline conditions

Question 29

Nomenclature for polynucleotides

Answer 29

* mononucleotide - 1 nucleotide (ATP, ADP, FMN) * dinucleotide - 2 nucleotides (FAD, NAD+, dCdG) * Tri-/Tetra-, etc * oligonucleotide (more than 20 <50 nucleotides) * polynucleotides (large number)

Question 30

Explain why the bases in a nucleic acid "stack: and identify the forces that favour stacking

Question 31

State Chargaff's rules and explain what they tell you about a nucleic acid

Answer 31

* nucleotides in DNA were not all present in equal numbers and base composition varies from organism to organism * Chargaff noted that the amount of A is equal to the amount of T, the amount of C is equal to the amount of G, and the total amount of A/G is equal to the total amount of C/T. * Chargaff’s “rules” could be satisfied by a molecule with two polynucleotide strands in which A and C in one strand pair with T and G in the other

Question 32

describe the double helical structure of B-DNA (6) ## Footnote B-form is the most common form of DNA

Answer 32

1. double helix: 2 chains connected to the other through H-bonds between bases (non-covalent), the strands are *anti-parallel*, with an overall right-handed twist 2. hydrophobic core/polar exterior - base pairs largely excluded from water (stabilizes H-bonds) and stack on top of each other - base pair edges are exposed in the major/minor grooves 3. The twisting of the DNA “ladder” into a helix creates two grooves of unequal width, the major and minor grooves. 4. ribose/deoxyribose and phosphates (very polar) exposed to water 5. H-bonds in pairing interactions 6. ~10 base pairs/turn ## Footnote antiparallel: complenatary strands should run in opposite directions of 5'-3'

Question 33

State how strands of DNA align with one another specifically rather than randomly

Answer 33

**hydrogen bonding between the four bases**: amino and carbonyl functional groups found in DNA (along with the right nitrogen atoms) allow for specific hydrogen bonding interactions between the bases. * 2 hydrogen bonds link A/T, T/A * 3 hydrogen bonds link G/C, C/G formation of base pairs requires the correct hydrogen bonding interactions between bases in the two strands

Question 34

Describe how the structure of double-stranded DNA is stabilized

Answer 34

1. **Hydrogen bonds** - between base pairs 2. **Base stacking (primary stabilising force)** - primarily van der Waals forces, hydrophobic forces

Question 35

State why it is important that the structure of DNA is stabilized by non-covalent forces only

Answer 35

The polar sugar/phosphate backbone is found on the outside while the more hydrophobic bases stack in the core of the structure

Question 36

State the role for the grooves in the structure of double-stranded DNA

Answer 36

- Other hydrogen bonding sites on the bases are open to the major and minor grooves where they can interact with solvent

Question 37

List the major differences between RNA and DNA 3-D structures

Answer 37

**DNA** - *double stranded* (helical), base pairs A/**T**, G/C, *inter*strand base pairing **RNA** - *single stranded*, base pairs A/**U**, G/C, *intra*strand base pairing, partially SS and partially duplex - much more dependant on nucleotide sequence than dsDNA

Question 38

State why the denaturation of a double helix is feasible and identify the chemical conditions that can cause denaturation

Answer 38

* denaturation must happen for replication or transcription * Increasing temperature

Question 39

Define "melting" of a double stranded molecule of DNA ## Footnote page 28

Answer 39

* separation of the two paired strands (dd→ss) * disruption of non-covalent forces (base-stacking/hydrogen bonding) * cooperative process: SLOW & hard to start, easy to continue (FAST zippering) * changes in absorption properties occur (increase in absorption at 260nm as strands separate * essential for some cellular processes (replication/transcription)

Question 40

Define the terms hyperchromicity and hypochromicity with respect to double-stranded nucleic acids

Answer 40

**hyperchromicity** - a relatively high absorbance, DS→SS DNA **hypochromicity** - low absorbance, SS→DS DNA

Question 41

Explain the term renaturation

Answer 41

* Reformation of DS DNA so it regains its native conformation * occurs in slow cooling (20-25℃ below Tₘ) - rapid coolin can result in improper base pairing 1. proper base pairing 2. cooperative: nucleation (slow), zippering (fast)

Question 42

what is Tₘ?

Answer 42

Tₘ - the midpoint of melting affected by sequence of nucleotides, composition, and solvent

Question 43

Describe and explain the relationship between the Tm of a double-stranded nucleic acid and its relative content of C and G

Answer 43

Tₘ is higher for for nucleic acids containing more C/G pairs * base stacking is stronger in G/C pairs: makes 3 hydrogen bonds → G donates 2 and accepts 1, C accepts 2 and donates 1 * A/G pairs only have 2 hydrogen bonds between them (regions of A/T pairs in DNA tend to denature first)

Question 44

State how and why the Tm of a nucleic acid is affected by pH and ionic strength ## Footnote slide 31

Answer 44

pH: increasing/decreasing pH will decrease Tm by affecting protonation state * Tm is highest at pH 7 salt concentrations: ions shield negative charges on phosphate backbone (Mg2+, positively charged proteins) * low salt → decreases Tm (destabilizes dbl helix) * high salt → increases Tm (stabilizes dbl helix)

Question 45

State whether or not RNA can "melt" and justify your answer

Answer 45

RNA can melt but it's ,=melting curve is less steep and starts at a higher absorbance due to having a mix of SS and DS RNA * A260nm of free nucleotides > SS > DS * Abs increases as disorder of structure increases