BIOC 2300 > nucleic acid structure + function - chp3 > Flashcards

nucleic acid structure + function - chp3 Flashcards

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1
Q

list three reasons why ATP hydrolysis is energy-favourable at physiologic conditions

A
  1. electrostatic repulsion : repulsion between charged phosphoryl groups destabilizes ATP (makes bonds want to break)
  2. resonance stabilization : the released phosphate ion has more possible resonance forms when covalently attached
  3. solvation / hydration : the phosphate ion & ADP have a greater degree of solvation and form hydration layers, making them more stable than ATP (water can surround them better now that they are separate)
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2
Q

how does resonance affect the stability of free phosphate ions?

A

the released phosphate ion has more possible resonance forms than when it is attached to adenylate, which favours the free phosphate ions over ATP or ADP

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3
Q

what are the three components that make up the structure of ATP?

A

adenine base, ribose sugar & three phosphoryl groups

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4
Q

what specific mutation causes Hutchinson-Gilford progeria syndrome?

A

a single point mutation, changing a Cytosine to a Thymine

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5
Q

how does the C1824T mutation affect the mRNA transcript of the lamin A gene?

A

it creates a new internal (cryptic) splice site in Exon 11. this leads to altered RNA splicing and a truncated exon 11 (splicing is a key feature in eukaryotes)

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6
Q

why is the toxic truncated protein (progerin) produce in progeria toxic to the cell?

A

cleavage does not occur, meaning the ICMT modification is not removed. this toxic protein causes the nuclear envelope to deform (no post-transcriptional modification on the gene)

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7
Q

how can you tell the difference between a ribonucleotide and a deoxyribonucleotide looking at the sugar?

A

look at the 2’ carbon of the ribose sugar:
- ribonucleotides : have a hydroxyl group in 2’ position
- deoxyribonucleotides have a hydrogen atom at the 2’ position

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8
Q

what type of bond links nucleotides together to form the backbone of DNA and RNA?

A

a phosphodiester bond

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9
Q

how and what direction are the nucleotides in RNA and DNA linked?

A

they are linked together through a phosphate backbone, in a linear direction

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10
Q

four rules to be considered an aromatic ring

A
  1. cyclic
  2. conjugated pi bonds
  3. planar (flat) - due to pi bonds
  4. Huchels rule (4n + 2 electrons)
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11
Q

describe the difference between major + minor grooves in DNA

A

these are indentations in the double helix, that arise from the way the strands are wound. the MAJOR groove is wider and more accessible for protein binding, compared to the MINOR groove

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12
Q

what is the primary structure?

A

the unique arrangement or sequence of deoxyribonucleotides (in DNA) or ribonucleotides (in RNA) arranged in a single chain, usually depicted as single letters in a row

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13
Q

what defines the secondary structure of DNA?

A

two complementary strands of DNA binding or annealing together in an antiparallel fashion to form a double helix

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14
Q

what rule states that the amount of A = T, and the amount of C = G?

A

chargaff’s rule

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15
Q

what are the specific base pairs formed by hydrogen bonding in DNA commonly called?

A

watson-crick base pairs

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16
Q

how many hydrogen bonds form between 1. A-T and 2. G-C

A
  1. two hydrogen bonds
  2. three hydrogen bonds
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17
Q

T/F base pairing occurs through the specific arrangement of hydrogen bond donors and acceptors on nitrogenous bases

A

true

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18
Q

how are nitrogenous bases oriented within the DNA double helix to allow for hydrogen bonding?

A

they are arranged in a planar fashion, parallel to adjacent bases on the same strand, and located in the interior of the helix

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19
Q

what specific type of interactions contribute to the stability provided by base stacking?

A

van der waals interactions (specifically when bases are at their van der waals distance) and hydrophobic effects

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20
Q

how are the base pairs oriented?

A

they are stacked upon each other within the interior of the helix, within van der waals distance

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21
Q

T/F hydrogen bonding contributes majorly to the overall stability of the helix

A

false : most of the stability of a DNA double helix is provided by base stacking

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22
Q

what is the hyperchromic shift in nucleic acid biochemistry?

A

it is the increase in absorbance at 260 nm that occurs when double-stranded DNA is denatured into single-stranded DNA

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23
Q

why does single-stranded DNA absorb more light than double-stranded DNA?

A

absorbance increases due to the exposure of nitrogenous bases, which were previously “buried” and stacked in the double helix

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24
Q

at what specific wavelength do nucleic acids (DNA and RNA) absorb light most strongly?

A

260 nm, this makes it super useful for identifying and quantifying them

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25
how is the hyperchromic shift practically used in a laboratory setting?
it is used to calculate the concentration of DNA and RNA based on Beer-Lambert law
26
what are the two alternative terms for DNA "denaturation" and "renaturation"
denaturation is also called melting, renaturation is also called annealing
27
what are the three conditions that can cause DNA denaturation?
heating, addition of acid, addition of base
28
define DNA denaturation at the structural level
the separation of the double helix into two individual strands
29
define DNA reformation at the structural level
the process where two individual DNA strands reform a helix
30
what does Tm represent on a DNA melting curve?
the melting temperature, which is the temperature at which 50% of the DNA is in a single-stranded state
31
what are the three primary differences between DNA and RNA?
sugar - DNA uses deoxyribose, RNA uses ribose base - DNA uses thymine, RNA uses uracil structure - DNA is typically a double helix, where RNA forms complex intrastrand structure
32
what is a ribozyme?
an RNA molecule that has catalytic activity, meaning it can act like an enzyme (DNA unable to form catalytic molecules)
33
provide an example of a ribozyme
RNAse P, which cleaves nucleic acids
34
T/F RNA is always a simple, single-stranded linear molecule
false, RNA can form complex intrastrand structures (secondary and tertiary structures) by folding back on itself
35
why is RNA most susceptible to spontaneous degradation than DNA?
the 2' OH group in RNA facilitates spontaneous degradation , which makes RNA less stable, as well as the ribose are not flat rings (aka planar) so no ring stacking
36
what are the three postulated methods of DNA replication?
- semi conservative - conservative - dispersive
37
define semi-conservative replication
each daughter DNA molecule contains one strand from the original template and one newly synthesized strand, discovered by Meselson and Stahl
38
what enzyme is responsible for catalyzing the addition of deoxyribonucleotides to a growing DNA strand?
DNA polymerase
39
in what direction does DNA polymerase synthesize a new DNA strand?
it adds nucleotides to the 3' end, meaning synthesis occurs in the 5' to 3' direction
40
what are the two types of strands formed at the replication fork?
leading strand : synthesized continuously lagging strand : synthesized discountinuously
41
what is required for the DNA polymerase to being synthesizing a new DNA strand?
primers
42
what is the correlation between caregiving / perceived stress and telomere length?
there is a negative correlation, increased chronicity of caregiving and higher perceived stress are associated with shorter telomere length
43
what is the function of the enzyme Telomerase?
it uses an internal RNA templates to synthesize a complementary strand of DNA in a 5' to 3' direction, extending to the ends of chromosomes
44
how does Telomerase bind to DNA?
through base pairing between the telomerase RNA template and the DNA
45
what role does Primase play in the telomere extension process?
it adds a new RNA primer to the extended DNA strand to facilitate further synthesis by the DNA polymerase complex
46
what enzyme is responsible for repairing the "gap" after the RNA primer is removed?
DNA ligase
47
what is the basic definition of DNA transcription?
one strand of DNA is used as a template for the synthesis of a complementary RNA strand
48
in what direction does mRNA synthesis occur?
in the 5' to 3' direction
49
how do the bases in the template DNA strand compared to the synthesized RNA strand?
the template DNA contains the complementary sequence to the RNA strand, with T in DNA replaced by U in RNA
50
what enzyme is responsible for synthesizing RNA, and what does it recognize to start the process?
RNA polymerase, it recognizes promoter sequences located upstream of the gene
51
what is the difference between the coding strand and template strand during transcription?
template strand : the DNA strand actually read by RNA polymerase to build the RNA coding strand : the DNA strand whose sequence matches the RNA except T/U, which is displaced during the process
52
what are the key differences in the timing and location of transcription and translation between prokaryotes and eukaryotes?
prokaryotes : transcription and translation occur simultaneously in the cytoplasm eukaryotes : transcription and processing happen in the nucleus, while translation occurs later in the cytoplasm
53
what specific mRNA sequence helps orient the ribosome in prokaryotes?
the Shine-Dalgarno sequence, which appears 5-13 base pairs upstream of the first AUG (start - MET) codon
54
how is protein translation terminated?
it is triggered by stop codon on the mRNA a release factor binds to the ribosome instead of a charged tRNA, causing the polypeptide chain to be released
55
describe the role of the spliceosome in eukaryotic cells
it is the machinery responsible for RNA processing, specifically in removing introns and joining exons together
56
what is the primary purpose of the polymerase chain reaction?
it is a method used to exponentially amplify a specific targeted DNA segment
57
name the three temperature phases of a PCR cycle in order
phase 1. DNA denaturation (separation of the two strands) phase 2. annealing (primer binding to the template) phase 3. primer extension and DNA synthesis (DNA polymerase makes the new DNA)
58
by what factor does the number of DNA molecules increase in each PCR cycle?
2n
59
taq DNA polymerase
extracted from thermophiles (high heat) to ensure they won't denature in the presence of heat from the PCR reaction
60
why is the temperature lowered during the second phase (annealing) of PCR
to promote primer annealing (the binding of primers P1 and P2 to the target DNA)
61
what is molecular cloning?
the creation of recombinant DNA, an essential process used to amplify and manipulate genes of interest
62
what are the four basic steps of molecular cloning workflow?
1. isolation of target DNA fragments (inserts) via PCR or restriction 2. ligation of inserts into a cloning vector (aka a plasmid) 3. transformation of the recombinant plasmid into a host (like bacteria - as much easier for bacteria to maintain) 4. selection of the hosts that contain the intended recombinant plasmid - as not all bacteria will be accepting it
63
what is the function of a restriction enzymes in cloning?
they are used to cut both the target DNA fragment and the vector to prepare them for joining
64
what enzyme is used to paste the digested DNA fragment into the vector?
DNA ligase
65
what is an MCS in the context of a cloning vector?
multiple cloning site (aka polylinker) which is a short segment of DNA that contains many restriction sites for easy insertion of DNA
66
what is the primary use of examining an individual's DNA sequence via Sanger sequencing?
it is useful for disease diagnosis, and in some cases, can even guide disease treatment
67
what is the key chemical component that makes Sanger sequencing possible?
the use of dideoxynucleotide triphosphates (ddNTPs) in a primer dependant DNA synthesis reaction
68
why does the incorporation of a ddNTP stop DNA synthesis?
because ddNTPs lack a 3' reactive hydroxyl group, which is necessary for forming a phosphodiester bond with the next nucleotide, thereby blocking elongation and terminating the reaction at random
69
how do bacteria protect themselves against invading viruses?
they protect themselves by degrading the viral DNA in two mechanisms 1. restriction endonuclease / enzyme that recognize ~6 bp palindromic sequences ("innate immunity") 2. CRISPR-CAS9 systems that recognize ~20 nucleotides ("adaptive immunity")
70
what is CRISPR and CAS?
CRISPR : clustered regularly interspersed short palindromic repeats (where bacteria collects the viral DNA, will remember and be passed down to their progeny) CAS : CRISPR-associated endonuclease (will cut double stranded DNA)
71
Streptoccocus thermophilus
showed that fermenting bacteria could be programmed to be resistant to a specific virus causing spoilage in dairy products, through the incorporation of that viral DNA sequence into a CRISPR region
72
CRISPR RNA
complements to the viral DNA recognition (will base pair with virus, aka target DNA)
73
tracrRNA
binds to CRISPR RNA and recruits CAS9, also base pair with CRISPR RNA
74
PAM
protospacer adjacent motif : how bacteria distinguishes from self vs viral DNA (don't have PAM, not going to cut)
75
why does it depend if CRISPR-CAS9 will help progeria?
it depends on timing, when the mutation was identified : can be too late after a child is already born, as the mutation is already in place and the damage is already done, can also happen so frequently within the DNA and CRISPR can also recognize ~20 nucleotides best place is during IVF, a fertilized egg
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