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MMBIO 240 > Exam 3 Review > Flashcards

Exam 3 Review Flashcards

1
Q

Define semiconservative replication

A
  • Double-stranded DNA molecule unwinds
  • Each strand serves as a template for the synthesis of a new complementary strand
  • Two resulting daughter DNA molecules
  • Each consists of one original parental strand and one newly synthesized strand
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2
Q

Explain the directionality of DNA synthesis. What is bidirectional replication? Why is a primer needed any time DNA synthesis occurs?

A

DNA synthesis proceeds in the 5’ -> 3’ direction
(new nucleotides are added to the growing 3’ OH end of existing strand)

Bidirectional Replication refers to DNA synthesis proceeding simultaneously in both directions (leading strand & lagging strand)

DNA polymerases cannot start synthesizing a new DNA strand without a short RNA or DNA PRIMER to provide the initial 3’ hydroxyl group to which new nucleotides can be added.

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3
Q

On a DNA replication diagram, be able to label (and explain) the following: replication fork, leading strand, lagging strand, RNA primer, Okazaki fragment, β clamp, DNA polymerase core, helicase, topoisomerase.

A
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4
Q

In DNA replication, why is the lagging strand fragmented, but the leading strand is continuous?

A
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5
Q

DNA polymerase has 3’-5’ exonuclease proofreading activity; what does this mean?

A
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6
Q

What are the functions of the following proteins: DNAB helicase, DNAG primase, DNA polymerase 1, DNA polymerase 3, single stranded bind proteins, DNA ligase

A
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7
Q

Describe the process of DNA replication on the leading strand.

A
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8
Q

Describe the process of DNA replication on the lagging strand including the mechanism of forming and repairing okazaki fragments.

A
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9
Q

What are the eukaryotic equivalents to the proteins used in bacterial DNA replication.

A
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10
Q

What is the purpose of the beta clamp in DNA synthesis?

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11
Q

What are the differences between the bacterial and eukaryotic holoenzymes?

A
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12
Q

Describe the steps in DNA synthesis and the proteins involved at each step.

A
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13
Q

Why do we have telomeres? What causes the need for telomeres? How does telomerase fix this?

A
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14
Q

What kind of polymerase is telomerase? (i.e., what does it read, and what does it make?)

A
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15
Q

Describe the types of mutations.

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16
Q

Why are most mutations benign?

A
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17
Q

Know the following enzymes of eukaryotic DNA replication. What are their functions? Were any alternative names for them mentioned in lecture?

A

Topoisomerase I

Helicase

DNA Pol Alpha

DNA Pol Delta

DNA Pol Epsilon

PCNA

Flap endonuclease I (FenI)

DNA Ligase I

Telomerase

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18
Q

What is a crossover event and during which part of cell division does it occur? What are recombinant chromatids and what is the chiasma?

A

Crossover event: Genetic exchange between homologous chromosomes
- occurs during meiosis in prophase I

Recombinant chromatids: Chromatids with exchanged genetic material
- result of crossing over

Chiasma: Site of crossing over between chromatids during meiosis
- x-shape made by the legs of 2 different sister chromatids

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19
Q

Characterize a double Holliday junction and what proteins help create the Holliday intermediate.

A

Double Holliday junction: Interlocked DNA structure formed during genetic recombination

Proteins involved: Enzymes like RecA, Rad51, and RuvC mediate Holliday junction formation

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20
Q

What is the difference between a holliday junction and a double holliday junction?

A

Holliday Junction:
- Involves two DNA double helices.
- Has four arms, each representing a DNA strand.
- Formed by a single crossover event.

Double Holliday Junction:
- Involves two Holliday junctions.
- Contains eight arms, each representing a DNA strand
- Formed by two crossover events.

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21
Q

Describe the steps involved in the process of crossing over and the proteins that aid this process?

A

STEPS INVOLVED
1. Alignment: Homologous chromosomes pair up during prophase I of meiosis.
2. Breakage: DNA breaks occur at corresponding positions on paired chromosomes.
3. Exchange: Broken ends of DNA strands swap segments with each other.
4. Rejoining: DNA strands recombine with their respective partners, forming recombinant chromatids.

PROTEINS INVOLVED
- RecA/Rad51: Catalyze strand invasion, facilitating DNA exchange.
- RuvABC: Resolves Holliday junctions, aiding in crossover formation.
- Topoisomerases: Relieve torsional strain, allowing DNA strands to unwind and recombine.
- Exonucleases: Trim and process DNA ends for proper recombination.

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22
Q

Define the following: bivalent, sister chromatid, recombinant chromatid, RuvA,B,C, RecA,B,C,D and RecFor, UVR

A
  • Bivalent: Pair of homologous chromosomes joined together during prophase I of meiosis.
  • Sister chromatid: Identical copies of a chromosome produced during DNA replication, held together by a centromere.
  • Recombinant chromatid: Chromatid resulting from the exchange of genetic material between non-sister chromatids during crossing over.
  • RuvA, RuvB, RuvC: Proteins involved in resolving Holliday junctions during genetic recombination (A&B help move DNA and C cleaves it).
  • RecA, RecB, RecC, RecD: Proteins involved in DNA repair, recombination, and regulation.
  • RecFOR: Protein complex involved in DNA recombination and repair.
  • UVR: Enzyme involved in nucleotide excision repair of DNA damaged by ultraviolet radiation.
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23
Q

What are the differences between Endonuclease and
Exonuclease?

A

Endonuclease: Cuts DNA or RNA at specific internal sites.
- Involved in processes like DNA cleavage during recombination and repair.
- Acts on specific sites

Exonuclease: Removes nucleotides from the ends of DNA or RNA strands.
- Involved in processes like proofreading during DNA replication and repair.
- Acts without specificity

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24
Q

What is the nascent strand?

A

The newly synthesized DNA strand during DNA replication. It is complementary to the template strand and is actively being synthesized by DNA polymerase.

25
Q

What are DNA dependent DNA polymerase (and other versions of this naming system)?

A

Enzymes that catalyze the synthesis of DNA using a DNA template
- RNA-dependent DNA polymerase (ex: reverse transcriptase)
- DNA-dependent RNA polymerase (used for normal transcription of RNA)
- RNA-dependent RNA polymerase (commonly found in viruses)
- DNA polymerase III (prokaryotic DNA replication)
- DNA polymerase alpha, delta, and epsilon (eukaryotic DNA polymerases involved in various aspects of DNA replication and repair)

26
Q

What are Hybrid strands?

A

DNA molecules or segments that contain both DNA and RNA nucleotides. These molecules are formed through processes like RNA-DNA hybridization, where single-stranded RNA binds to complementary single-stranded DNA to form a hybrid duplex.

27
Q

What is Processivity?

A

The ability of an enzyme to catalyze multiple sequential reactions without dissociating from its substrate

28
Q

What is a Holoenzyme?

A

Complete, active form of an enzyme, consisting of both protein and non-protein components (cofactors or coenzymes).

29
Q

What are rNTPs?
What are dNTPs?

A

rNTPs: Ribonucleotide triphosphates, used as building blocks for RNA synthesis during transcription.

dNTPs: Deoxyribonucleotide triphosphates, used as building blocks for DNA synthesis during replication and repair.

30
Q

What is a Clamp loader?

A

Protein complex that loads sliding clamps onto DNA strands during DNA replication to increase processivity of DNA polymerases

31
Q

Describe Elongation,
Polymerization, and
Ligation

A

Elongation: The phase of DNA replication or transcription where nucleotides are added to a growing strand.

Polymerization: The process of forming a polymer, such as joining nucleotides to form a DNA or RNA strand.

Ligation: The process of joining two DNA or RNA fragments together by forming a phosphodiester bond between them

32
Q

What are the differences between these mutations?

Nonmutant
Silent mutation
Missense mutation
(Stop mutation)
Synonymous mutation
(Nonsense mutation)
Nonsynonymous mutation
Frameshift mutation
Point mutation

A

Nonmutant: Refers to the standard or wild-type version of a gene without any mutations.

Silent mutation: A mutation that does not change the amino acid sequence of a protein due to redundancy in the genetic code.

Nonsense mutation: A mutation that introduces a premature stop codon, leading to truncated or non-functional proteins (a.k.a. stop mutation)

Synonymous mutation: A mutation that changes a nucleotide in the DNA sequence but does not change the amino acid sequence of the protein.

Nonsynonymous mutation: A mutation that changes a nucleotide in the DNA sequence, resulting in a different amino acid in the protein (a.k.a. missense).

Frameshift mutation: A mutation that inserts or deletes 1, 2, or 4 nucleotides in a DNA sequence, causing a shift in the reading frame during translation.

Point mutation: A mutation that affects a single nucleotide base in a DNA sequence.

33
Q

Describe each of these:

Insertion
Deletion
Translocation

A

Insertion: Addition of one or more nucleotides into a DNA sequence.

Deletion: Removal of one or more nucleotides from a DNA sequence.

Translocation: Movement of a segment of DNA from one chromosome to another, often leading to genetic abnormalities.

34
Q

What is a Fusion gene?

A

Fusion gene: A hybrid gene formed by the joining of two separate genes, often as a result of chromosomal rearrangements such as translocations.

35
Q

What is an Indel?

A

Indel: Short for “insertion-deletion”, refers to a mutation in DNA involving the insertion or deletion of one or more nucleotides, leading to alterations in the genetic sequence.

36
Q

What is a Codon?

A

Codon: A sequence of three nucleotides in DNA or RNA that codes for a specific amino acid or serves as a start or stop signal for protein synthesis.

37
Q

What is a Reading frame?

A

Reading frame: The specific way in which a sequence of nucleotides is read during translation to determine the sequence of amino acids in a protein. It starts at a specific codon (usually AUG) and proceeds in triplets along the mRNA molecule.

38
Q

What is a Oncogene?

A

Oncogene: A gene that has the potential to cause cancer when mutated or overexpressed, typically by promoting cell proliferation or inhibiting apoptosis.

39
Q

That is a Tumor suppressor gene?

A

Tumor suppressor gene: A gene that helps regulate cell growth and division, and when mutated or inactivated, can contribute to the development of cancer by allowing uncontrolled cell growth.

40
Q

Name and describe each of the following systems

MMR system
BER system
NER system

A

MMR system: Mismatch repair system, responsible for correcting errors such as base mismatches and small insertions/deletions that occur during DNA replication.

BER system: Base excision repair system, which repairs damaged DNA bases by removing the damaged base and replacing it with a correct one.

NER system: Nucleotide excision repair system, which repairs bulky DNA lesions such as those caused by UV radiation or certain chemicals by removing the damaged section of DNA and replacing it with new nucleotides.

41
Q

What is the Abasic Site (a.k.a. AP Site)?

A

Abasic site: A site in DNA where the nitrogenous base is missing due to hydrolysis or chemical damage, often resulting from spontaneous depurination or exposure to certain chemicals or radiation. It is also known as an AP site (apurinic/apyrimidinic site).

42
Q

Describe Upstream
Describe Downstream

A

Upstream: Refers to the direction towards the 5’ end of a nucleic acid sequence, typically the direction opposite to that of transcription or replication.

Downstream: Refers to the direction towards the 3’ end of a nucleic acid sequence, typically the direction of transcription or replication.

43
Q

How are double-stranded breaks repaired?

A

TWO MECHANISMS:

Non-homologous end joining (NHEJ)

Homologous end joining (HEJ) or Homologous repair (HR)

44
Q

What is RecA and what is its function?

A

Protein involved in DNA repair and recombination in bacteria. Its main function is to facilitate homologous recombination, particularly during the repair of DNA double-strand breaks.

  • The RecA protein plays its central role by binding single-stranded DNA (ssDNA) to form a presynaptic filament that searches for a homologous double-stranded DNA (dsDNA) donor from which to repair
  • RecA promotes the exchange of DNA strands between homologous regions, allowing for the repair of damaged DNA and the generation of genetic diversity through genetic recombination.
45
Q

What is NHEJ? Why is it significant? Compare and contrast with normal double-stranded break repair.

A

NHEJ (Non-homologous end joining):
- Joins broken DNA ends directly.
- Can be error-prone, leading to insertions or deletions.
- Significant for its rapid response to repair DNA breaks but may introduce mutations.

Normal HEJ (homologous end joining) repair includes HR (homologous recombination):
- HR uses homologous template for accurate repair, occurring in S/G2 phases.
- HR maintains genomic integrity, whereas NHEJ may introduce mutations.

46
Q

Be able to diagram/describe HEJ and NHEJ

A

BOTH REPAIR Double-Stranded Breaks In DNA

HEJ: Homologous End Joining (requires homologous sequence to guide repair)

NHEJ: Non-Homologous End Joining (directly ligated without needing homologous template)

47
Q

What proteins are involved in NHEJ and HEJ? What are their functions.
Ex. What would happen if a fatal mutation occurred in Artemis?

A

??

In non-homologous end joining (NHEJ):

Ku70/Ku80 heterodimer: Recognizes and binds to broken DNA ends, facilitating their alignment.
DNA-PKcs (DNA-dependent protein kinase catalytic subunit): Phosphorylates nearby proteins and activates downstream repair factors.
XRCC4 (X-ray repair cross-complementing protein 4): Forms a complex with DNA ligase IV, stabilizing the ligation process.
DNA ligase IV: Catalyzes the final step of joining the broken DNA ends together.
In homologous recombination (HR), often referred to as HEJ:

Rad51: Catalyzes strand invasion, facilitating homology search and DNA strand exchange.
BRCA1/BRCA2 (Breast cancer type 1/2 susceptibility proteins): Facilitate Rad51 function and regulate HR.
Rad52: Promotes annealing of complementary DNA strands.
Rad54: Stimulates Rad51-mediated DNA strand exchange and recombination.
If a fatal mutation occurred in Artemis:

Artemis deficiency disrupts the NHEJ pathway.
Impaired DNA repair of double-strand breaks.
Accumulation of unrepaired DNA damage.
Increased genomic instability and susceptibility to cancer.
Severe combined immunodeficiency (SCID) due to impaired V(D)J recombination, leading to compromised immune function.

48
Q

What are transposons?

A

Transposons, also known as “jumping genes,” are DNA sequences that can move or transpose themselves to different positions within the genome. They are capable of changing their genomic location, potentially causing mutations, altering gene expression, or contributing to genetic diversity.

49
Q

What are the three major kinds of transposons discussed in lecture?

A

DNA transposons: These transposons move directly by excising themselves from one genomic location and inserting into another. They typically encode a transposase enzyme responsible for catalyzing their movement.
Retrotransposons: These transposons move via an RNA intermediate. They are first transcribed into RNA, reverse transcribed into DNA by a reverse transcriptase enzyme, and then inserted into a new genomic location. Retrotransposons include LTR retrotransposons (long terminal repeat retrotransposons) and non-LTR retrotransposons.
LTR retrotransposons: These are retrotransposons that contain long terminal repeats (LTRs) at their ends, similar to retroviruses. They encode reverse transcriptase and integrase enzymes necessary for their mobility.

50
Q

What is transcription? How does it fit into the central dogma?

A

DNA
->Transcription
RNA
-> Translation
Protein

51
Q

What are the three types of RNA? What are the characteristics of each?

A

Messenger RNA (mRNA):
- Carries genetic information from DNA in the nucleus to the ribosome in the cytoplasm.
- Contains codons that specify the sequence of amino acids in a protein.
- Generally unstable and short-lived.

Ribosomal RNA (rRNA):
- Integral part of ribosomes, the cellular machinery responsible for protein synthesis.
- Combines with proteins to form ribosomal subunits.
- Catalyzes the formation of peptide bonds between amino acids during translation.

Transfer RNA (tRNA):
- Carries specific amino acids to the ribosome during protein synthesis.
- Contains an anticodon that base-pairs with the corresponding codon on mRNA.
- Adopts a cloverleaf-like secondary structure with three loops and an amino acid attachment site at one end.

52
Q

What are the major subunits of the bacterial RNA polymerase core? What is the function of the sigma factor, and why is it different from the core subunits?

A

The major subunits of the bacterial RNA polymerase core are:

α (alpha)
β (beta)
β’ (beta prime)
ω (omega)
The function of the sigma factor is to recognize and bind to specific DNA sequences called promoters, which initiate transcription. The sigma factor guides the core RNA polymerase to the promoter region and helps initiate transcription by facilitating the unwinding of DNA and the synthesis of the initial RNA transcript.

The sigma factor is different from the core subunits because it is not part of the catalytic center responsible for RNA synthesis. Instead, it acts as a regulatory protein that associates with the core enzyme to direct it to specific DNA sequences where transcription should begin. Once transcription initiation occurs, the sigma factor dissociates from the core enzyme, allowing elongation to proceed with the core enzyme alone.

53
Q

What are the two main types of transcription initiation discussed in lecture? What proteins are involved in each?

A
54
Q

What are the differences between kinetic and nucleolytic proofreading?

A
55
Q

What is rho (⍴) and what does it do during ⍴-dependent termination? What happens in ⍴-independent termination?

A
56
Q

What is DNA footprinting and why is it useful? What proteins are involved? How do you perform DNA footprinting?

A
57
Q

What is the function/purpose of an operon?

A
58
Q

What genes are encoded by the lac operon? What is the significance of each gene?

A
59
Q

How is the lac operon controlled?

A

MMBIO 240 (15 decks)

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