Nucleotides and Nucleic Acids Flashcards by Hannah Verbaas

What does it mean if a base is heterocyclic?

It has carbon and another atom within its ring

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What is the difference between purine and pyrimidine bases?

in terms of their structure

Purines have two carbon-nitrogen ring bases whereas pyrimidines have one carbon-nitrogen ring base.

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If there is a N with two single bonds, and a H (in an aromatic ring), is their lone pair is involved in resonance

The lone pair IS involved in resonance; the hydrogen can only be donated

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If there is a N with one single and one double bond (in an aromatic ring), is their lone pair in resonance?

The lone pair is NOT involved in resonance, it can only accept hydrogens

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If there is a N bonded to a C, which is double bonded to an O, is their lone pair involved in resonance?

The lone pair IS involved in resonance, it can only donate hydrogens

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If there is a N bonded to a C, which is double bonded to a N, is their lone pair involved in resonance?

The lone pair IS involved in resonance, they can only donate hydrogens (the single bonded N not the double bonded)

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What is a general trend for hydrogen bonding?

If the nitrogen is bonded with a hydrogen(s), it will donate

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What is the sugar portion of RNA?

Ribose

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What is RNA a polymer of?

G, A, C, U

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What is different about the structure of ribose’s ring?

C5’ is the only carbon not part of the ring structure

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what is the sugar portion of DNA?

Deoxyribose

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What differentiates ribose and deoxyribose?

Deoxyribose is missing an oxygen (OH to H) on 2’ C

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What is DNA a polymer of?

G, A, C, T

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What is a nucleoside?

Purine and pyrimidine bases linked to a pentose sugar

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How can you determine what carbon is being referred to in a nucleoside?

Carbons in the sugar ring are numbered from 1’ to 5’, which the carbons in the base are not prime

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Naming nucleosides

-ine in purines becomes…

osine

adenine + ribose = adenosine
guanine + ribose = guanosine

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Naming nucleosides

the ending in pyrimidines now end in…

idine

Cytosine + ribose = cytidine
Uracil + ribose = uridine

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where are sugars attached to bases in nucleotides?

N1 in pyrimidines, N9 in purines

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What are nucleotides?

Nucleosides + phosphate(s)

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what kind of reaction is the formation of a phosphoester bond?

Dehydration: the hydroxyl group loses a H, and the phosphate group loses OH

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What is a phosphoester bond?

A linkage between a phosphate and a hydroxyl group

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Hack to tell the difference between pyrimidine, and purine

Purine has only one nitrogen in its base ring

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What is a phosphodiester bond?

A linkage between two different carbons to one phosphate group

COPOC

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What type of reaction is the formation of a phosphodiester bond?

Dehydration

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What is the differences between: * Ester bonds * Phosphoester bonds * Phosphodiester bonds

* Ester = O=C - O * Phosphoester = COPO * Phosphodiester = COPOC

How can you identify a phosphoanhydride bond?

It POPs out at you

How are nucleotide sequences written out, in terms of their structures?

5' end (left) to 3' end (right ## Footnote AGTC which means the deoxyadenosine (mono/di/tri)phosphate is at the top/left

How do you determine the 5' carbon in a sequence?

You look for the 5'C not involved in a phosphodiester bond (not linked to another nucelotide)

What are two important characteristics of the DNA backbone?

It is polar and uniformly negatively charged (DNA is a negatively charged molecule)

Define residue

a single unit that makes up a polymer, such as an amino acid in a polypeptide or protein

What is the difference between the backbone in DNA vs. RNA?

Each residue in RNA has free 2' -OH groups

Why are the free 2' OH groups in RNA important?

They allow for extra hydrogen bonds to formed

How do you do sequence notation in DNA? | ex. 5'ATG3'

dAdTdG or ATG (5,3 notation is assumed)

How do you do sequence notation for RNA? | ex. 5'UCG3'

UCG

What does "p" mean in sequence notation?

Phosphorylated; indicates that it's linked via phosphodiester bonds

What is phosphorylated in this sequence? | pApCpGpCpT

The 5' end in phosphorylated

Nucleotides (NTP) become ____ in a polynucleotide change

Residues

How do you identify the 3' end of an RNA molecule?

It does not participate in a phosphodiester, 3'C is not linked to another nucleotide

What is the differentiation between oligo and polynucleotides?

* Oligo (general number) *20-50/60, not biological in nature* * Poly (large number) *very large, biological in nature (DNA, RNA)*

What is enzymatic hydrolysis of phosphodiester bonds?

DNase or RNase breaks (most commonly 3' side bond) to hydrolyze

Enzymatic Hydrolysis of Phosphodiester Bonds on the 5' side

What is alkaline hydrolysis of phosphodiester bonds?

Where the oxygen (5' side attached to phosphorus) become negative, and attacks the phosphorus which breaks the bond

What causes the ring in alkaline hydrolysis to break?

Steric stress

What happens to the RNA molecules that undergo alkaline hydrolysis?

* new molecules with no phosphate at the 5' end * 50%/50% of molecules with either the 2' or 3' ends phosphorylated

Why is alkaline hydrolysis "worse" than enzymatic?

It is not regulated, as its due to pH in environment (at pH 9) thus less ideal

When do bases best absorb?

~260nm

Are bases soluble or insoluble? Why?

They're poorly soluble due to their aromatic rings

Are bases hydrophobic or hydrophilic?

Hydrophobic with some polar groups

Is the sugar phosphate backbone non-polar or polar?

Polar

DNA absorbance is decreased by ____ % at 280nm compared to its absorbance at 260nm

50%

Why do the bases absorb ultraviolet light?

Aromatic rings which cause electron delocalization

When do proteins have the greatest absorbance?

280nm

How does someone test the potential purity of DNA? ## Footnote `

Abs. 260nm/ Abs. 280nm It should be 1.8-1.95

How does someone test the potential purity of RNA? ## Footnote `

Abs. 260nm/ Abs. 280nm It should be ~2.0

What is used to measure the concentration of nucleic acids in solution?

Absorbance at 260nm

What is the primary structure of nucelic acids?

Sequence of nucleotide residues

How are the primary structures stabilized?

Phosphodiester bonds

What are higher order structures of nucleic acids?

Chains of nucleotides

What type of protein structure is the DNA double helix?

Secondary

How are the chains of DNA connected? Ex. ss becoming ds

Hydrogen bonds between the bases

What is Chargaff's rule?

The ratio of G/C and A/T will be 50/50

What is the limitations of Chargaff's law? When is it applicable?

It is only true for dsDNA, as RNA and DNA are both singular strands thus can have a random composition on its chain

What kind of bond is between adenine and thymine?

Double

What type of bond is between Cytosine and Guanine?

Triple

Are hydrogen bonds stronger or weaker than covalent bonds? Why are they useful?

MUCH weaker, but they help stabilize the structure because of their large numbers

For H-bonds to form between or within biochemical macromolecules, they must be ____

shielded from water

When groups are hydrogen bonded to another, are they less or more polar?

They become less polar

If water is present in the formation of DNA, what happens?

It will not occur as water is "greedy" for hydrogen bonds, and thus the bonds wont form between the bases

What groups allow for hydrogen bonding interactions between bases in DNA?

Amino and carbonyl

What is the B-form of DNA?

'Common' DNA

What is the structure of common DNA?

Two strands that are anti-parallel, with an overall right-handed twist

What are the differences between the three forms of DNA

* A-DNA *dehydrated R-twist* * B-DNA *standard form* * Z-DNA *Left handed helix

What stabilizes dsDNA?

**MAJOR**: van der Waals and hydrophobic forces (only DNA, not other proteins) MINOR: base stacking interactions and hydrogen bonds

What stabilizes single strand DNA?

Covalent phosphodiester stabilizing bonds

Proteins can bind in the ____ between _______

grooves, two helical strands

What is the polarity of the core and backbone of DNA?

Hydrophobic core/ polar exterior

How is the hydrophobic core of DNA made?

Interactions shove the strands together, with water absent, hydrogen bonding of the bases then occurs

What parts of dsDNA are exposed to water?

Ribose/deoxyribose and phosphates (the backbone)

Axial view of DNA

Define complementarity

Ability to hydrogen bond with another anti parallel strand with hydrophobic interactions

What is the compliment of TAGTAG?

CTACTA not ATCATC, as it is read 5' to 3' but it is anti-parallel (correct answer will be "backwards" and not line up underneath the given sequence) *you are reading the answer as 5' to 3' so it doesnt matter than its "written" 3' to 5'*

Phosphates attached to a single C/OH groups are indentified using ____ prefixes

Mono/di/tri

Phosphates attached to multiple C/OH groups are described using ____ prefixes

Bis/tris

Out of dsDNA, ssDNA, and free nucleotides, order them in their absorbance at 260nm

free nucelotides> ssDNA > dsDNA

What is DNA melting?

the process by which double-stranded deoxyribonucleic acid unwinds and separates into single-stranded strands (denaturation)

At what temperature does DNA begin melting?

around 55 degrees

At what temperature does DNA melting end?

~80

How does DNA melting work?

H-bonds and base-stacking are weak forces which can be broken via heat

What is Tm?

The midpoint of melting; it also relates to stability (low Tm = low S)

Why does UV absorbance increase as DNA melts?

DNA unwinds to form single-stranded DNA. The bases become unstacked and can thus absorb more light.

What happens to absorbance if phosphodiester bonds are broken?

It'll increase however breaking these bonds are more difficult

DNA melting curves

## Footnote ssDNA has greater absorbance than dsDNA

Hyperchromic shift

Shift from low to high absorbance (ds to ss)

Hypochromic shift

Shift from high to low absorbance (ss to ds)

Hyperchromicity

A relatively high absorbance (ssDNA)

Process of denaturation

On what type of DNA can renaturation be done?

Plasmid; genomic is too long, and with its complexity, the process would be too slow

What is the temperature difference between denaturation and renaturation?

High for denaturation, low for renaturation

What are the steps of renaturation?

nucelation, and then zippering

# Define renaturation

Reformation of dsDNA so it regains its native conformation

What is an index of the thermal stability of a nucleic acid?

How does base number affect thermal stability?

greater base number (#)/length of the nucleic acid strands, greater the Tm

What base composition is more thermally stable? GC or AT

GC as its a triple bond

What do the melting curves for AT, regular DNA, and GC look like?

What base pair has stronger base stacking interactions?

Does DNA denature all at once or in regions?

In regions, AT rich regions will denature most readily

What is pKa?

An index used to express the strength of a (weak) acid. The smaller the pKa value, the stronger the acid

pKa values of the -OH groups will change if the _______ is part of a molecule

phosphate

Does more hydrogen in a molecule increase or decrease pKa?

More hydrogen decreases pKa

How does changing pH affect thermal stability?

It affects the protonation state of DNA and ability to H-bond

# Define Tm (thermal stability)

temperature to partially denature (50%), gives an idea about stability of the proteins

What molecules can form double-stranded helixes?

DNA and complementary RNA

What type of a helix is DNA?

Right-handed