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Biology 260 > Unit 5 Discussion Questions (Zoom) > Flashcards

Unit 5 Discussion Questions (Zoom) Flashcards

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1
Q

How is replication similar in bacteria & eukaryotes?

A

Similar:

Direction of replication: 5’ to 3’

Name of Enzymes needed:
- DNA polymerase (4)
- Primase (gyrase/helicase)
- Helicase
- DNA ligase
- Topoisomerase
- SSB

Anything else needed: RNA primer

Both have Lagging and leading strands

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2
Q

How is replication different in bacteria and eukaryotes?

A

Different
Location:
Prokaryotes: cytoplasm
Eukaryotes: nucleus, mitochondria, chloroplast

# of Origins:
Prokaryotes: 1
Eukaryotes: more than 1

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3
Q

How is transcription similar in bacteria & eukaryotes?

A

Both use RNA polymerase enzymes to synthesize RNA from a DNA template during transcription

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4
Q

How is transcription different in bacteria and eukaryotes?

A

Location
Prokaryotes: cytoplasm
Eukaryotes: nucleus, mitochondria, chloroplast

How it works:
Prokaryotes: more than 1 gene -> 1mRNA
Eukaryotes: 1 gene -> 1mRNA

Post transcriptional modification of mRNA:
Prokaryotes: None
Eukaryotes: 5’ cap, 3’ poly A tail, and slicing of introns

Types of RNA polymerases:
Prokaryotes: 1
Eukaryotes: 3

What binds RNA polymerase to promoter:
Prokaryotes: sigma factors
Eukaryotes: transcription factors

Are there operons:
Prokaryotes: Yes
Eukaryotes: No

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5
Q

How is translation similar in bacteria & eukaryotes?

A

Location:
Prokaryotes: cytoplasm
Eukaryotes: cytoplasm (+)

Direction mRNA is read:
5’ to 3’

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6
Q

How is translation different in bacteria and eukaryotes?

A

Location:
Eukaryotes also have it in RER, mitochondria and chloroplast

1 mRNA codes for how many proteins:
Prokaryotes: more than 1
Eukaryotes: 1

Type of ribosome:
Prokaryotes: 70s
Eukaryotes: 80s (except mitochondria)

1st Amino acid in polypeptide:
Prokaryotes: formyl-Methionine
Eukaryotes: Methionine

Can if occur while transcription is ongoing:
Prokaryotes: Yes
Eukaryotes: No

How does ribosome find the beginning of the mRNA:
Prokaryotes: Shine-Dalgarno sequence
Eukaryotes: 5’ cap initiation factors

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7
Q

Can we use differences between bacterial & eukaryotic processes of replication, transcription and/or translation to develop antibacterial drugs? Explain.

A

Yes, the differences in bacterial and eukaryotic processes of replication, transcription, and translation are key targets for antibacterial drugs;

For example, antibiotics like (rifampicin) target bacterial RNA polymerase without affecting eukaryotic polymerases

Drugs like (tetracyclines and macrolides) specifically bind to bacterial ribosomes, inhibiting protein synthesis while leaving eukaryotic ribosomes unaffected.

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8
Q

A scientist uses a molecule of DNA composed of nucleotides containing radioactive deoxyribose as a template for replication and transcription in a nonradioactive environment. What percentage of DNA strands will be radioactive after three DNA replication cycles? What percentage of RNA molecules will be radioactive?

A

(Start at 50%)

After 3 replication cycles 12.5% of the DNA strands will be radioactive (because replication is semiconservative)

0% of the RNA molecules will be radioactive (because transcription requires ribonucleotide not deoxyribonucleotide & only uses DNA as a template to make RNA)

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9
Q

What is semiconservative in regards to replication?

A

Semiconservative means that one strand of DNA is the parental and the newly synthesized one is complementary to it

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10
Q

Corynebacterium diphtheriae, the causative agent of diphtheria, secretes a toxin that enzymatically inactivates all molecules of elongation factor in a eukaryotic cell. What immediate and long-term effects does this have on cellular metabolism?

A
  • Elongation factors are used in translation

Inhibit elongation -> no new proteins can be made

  • No new proteins -> the cell will eventually lose its ability to maintain structures, repair damage, and conduct essential metabolic processes.
  • Proteins have a “shelf-life” so eventually existing proteins degrade
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11
Q

Hydrogen bonds between complementary nucleotides are crucial to the structure of dsDNA because they hold the two strands together. Why couldn’t the two strands be effectively linked by covalent bonds?

A

Remember that replication and transcription require that the two strands of DNA separate before these processes can occur

To break a covalent bond, requires lots of energy

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12
Q

On average, RNA polymerase makes one error for every 10,000 nucleotides it incorporates into RNA. In contrast, only one base pair error remains for every 10 billion nucleotides during DNA replication. Explain why the accuracy of RNA transcription is not as critical as the accuracy of DNA replication.

A

The accuracy of RNA transcription is less critical than DNA replication because RNA errors are temporary and only affect individual protein molecules

DNA replication errors are permanent and can be passed down to future generations, potentially causing harmful mutations

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13
Q

1) The lactose (lac) operon is an _________operon which is always ______.

A

The lactose (lac) operon is an inducible operon which is always off.

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14
Q

2) The regulatory gene produces a _______________ that binds to the DNA region of the operon called the _________________.

Once bound at this region, _________________________ is blocked from __________________ the genes of the operon thus no mRNA is made.

A

The regulatory gene produces a repressor that binds to the DNA region of the operon called the operator.

Once bound at this region, RNA polymerase is blocked from transcribing the genes of the operon thus no mRNA is made.

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15
Q

3) When lactose is present, the bacterium converts it to allolactose that binds to the ___________________ inactivating it by changing the 3D structure of the protein so that it can’t bind to the DNA anymore.

Now, __________________can bind to the promoter region and _____________________ can occur.

Ribosomes then translate the mRNA making the enzymes that
_____________lactose.

A

When lactose is present, the bacterium converts it to allolactose that binds to the repressor inactivating it by changing the 3D structure of the protein so that it can’t bind to the DNA anymore.

Now, RNA polymerase can bind to the promoter region and transcription can occur.

Ribosomes then translate the mRNA making the enzymes that catabolize lactose.

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16
Q

1) The tryptophan (trp) operon is a ____________operon that is always ________.

A

The tryptophan (trp) operon is a repressible operon that is always on.

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17
Q

2) In contrast to the lac operon, in the Trp operon the regulatory gene makes a __________________ but in this case this protein is inactive and can’t bind to the DNA region of the operon.

___________________ can bind to the promoter & mRNA is made.

A

In contrast to the lac operon, in the Trp operon the regulatory gene makes a repressor but in this case this protein is inactive and can’t bind to the DNA region of the operon.

RNA polymerase can bind to the promoter & mRNA is made.

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18
Q

3) The mRNA is translated by the cell’s free ribosomes into the enzymes that synthesize tryptophan. When tryptophan levels rise, tryptophan binds to the ____________________activating it by changing its 3D structure.

A

The mRNA is translated by the cell’s free ribosomes into the enzymes that synthesize tryptophan. When tryptophan levels rise, tryptophan binds to the repressor activating it by changing its 3D structure.

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19
Q

4) Now, the __________________can bind to the DNA region known as ________________ thus preventing ______________________ from binding to the promoter region.

__________________ no longer occurs.

A

Now, the repressor can bind to the DNA region known as the operator thus preventing RNA polymerase from binding to the promoter region.

Transcription no longer occurs.

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20
Q

If a mutation occurred in the regulatory gene of the lac operon producing a protein that doesn’t recognize lactose (allolactose) then what would be a likely outcome?

A

Can’t bind to lactose -> repressor is active and continues to bind on operator -> block RNA polymerase -> no transcription

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21
Q

If a stop codon is inserted into the middle of the regulatory gene of the trp operon producing a very short nonfunctional protein, then what would be a likely outcome?

A

Regulatory gene make repressor -> stop codon creates non-functional repressor -> repressor inactive so doesn’t bind to to operator -> continuous transcription

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22
Q

Lactose levels low, Glucose levels high

A

Operon off

  • No transcription; repressor active blocks operator
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23
Q

Lactose levels low, Glucose levels low

A

Operon off

-No transcription; repressor active blocks operator

-CAP active bound to promoter; RNA polymerase finds promoter

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24
Q

Lactose levels high, Glucose levels high

A

Operon off

  • No transcription; repressor inactive but CAP inactive not bound to promoter to aid RNA polymerase
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25
Q

Lactose levels high, Glucose levels low

A

Operon on

  • Transcription occurs; repressor inactive;
  • CAP active bound to promoter; RNA polymerase functions
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26
Q

Explain why an insertion of three nucleotides is less likely to result in a deleterious effect than an insertion of a single nucleotide.

A

Missense occurs when 3 nucleotides are added
3 new nucleotides just adds one amino acid to the chain

Inserting one base creates a frameshift
1 changes the whole chain because it shifts

27
Q

What is siRNA?

A

siRNA (small interfering RNA) is a double-stranded RNA molecule that mediates RNA interference by degrading specific mRNA molecules to prevent translation, thereby regulating gene expression.

28
Q

What is miRNA?

A

miRNA (microRNA) is a small, single-stranded RNA molecule that binds to complementary sequences on mRNA, usually resulting in translational repression or mRNA degradation, which fine-tunes gene expression.

29
Q

How could scientists use siRNA to turn off a cancer-inducing gene?

A
  • siRNA targets specific sequences in DNA
  • They can block creation or growth
  • Scientists can synthesize synthetic RNA
30
Q

Compare & contrast transduction, transformation, and conjugation

A

Similar all 3:
- Horizontal gene transfer
- Genetic diversity
- Change phenotype
- Can possibly incorporate DNA into host chromosome

Transformation:
Obtain DNA from the environment
Must be competent (able to do this)

Transduction:
Uses bacteriophage

Conjugation:
Uses pilus
Has fertility plasmid (F plasmid)

31
Q
  1. Describe how you think Biorad scientist made pGlo (based upon your knowledge of recombinant DNA technology.
  2. Then describe how your team proceeded to get the bacteria to take up this plasmid.
  3. In what way did you check to see if the bacteria were transformed?
  4. In what way did you check to see if the transformed bacteria were expressing green fluorescent protein (GFP)?
A
  1. Bio-Rad scientists likely made pGLO by using recombinant DNA technology to insert the green fluorescent protein (GFP) gene, along with an antibiotic resistance gene and a promoter, into a plasmid vector.
  2. To get the bacteria to take up the pGLO plasmid, our team treated the cells to make them competent, then performed a heat-shock transformation to encourage plasmid uptake.
  3. We checked if the bacteria were transformed by growing them on antibiotic-containing agar plates, as only transformed bacteria with the plasmid would survive.
  4. To verify if the transformed bacteria were expressing GFP, we observed the colonies under UV light, where successfully transformed bacteria would fluoresce green due to GFP expression.
32
Q

Suppose you want to insert into your dog a gene that encodes a protein that protects dogs from heartworms. A dog’s cells are not competent, so they cannot take up the gene from the environment; but you have a plasmid, a competent bacterium, and a related (though incompetent) F+ bacterium that lives as an intracellular parasite in dogs. Describe a possible scenario by which you could use natural processes to genetically alter your dog to be heartworm resistant.

A

To genetically alter the dog to be heartworm-resistant, you could insert the protective gene into a plasmid and transform the competent bacterium with this plasmid.

Then, you could allow the transformed bacterium to conjugate with the F+ bacterium that can live inside dog cells.

During conjugation, the protective gene would be transferred from the competent bacterium to the F+ intracellular bacterium.

Once this modified F+ bacterium carries the protective gene, it could be introduced into the dog’s cells.

Inside the dog, the F+ bacterium could continually express the protective protein, potentially providing the dog with resistance to heartworm infection.

33
Q

Draw the following TSI tubes with appropriate labels & colors:

No reaction (NR/NR)
Acid/Acid (A/A)
Alkaline/Acid (K/A)
Alkaline/Alkaline (K/K)
Alkaline/H2S Acid gas (K+H2S/A+H2S)
Alkaline/H2S (K/H2S)
Acid/H2S acid
Acid/Acid gas (A/A)

Identify which ones used lac operon

A
  • No reaction (NR/NR): All orange
  • Acid/Acid (A/A): All yellow
  • Alkaline/Acid (K/A): Red slant, yellow butt
  • Alkaline/Alkaline (K/K): All red
  • Alkaline/H2S Acid gas (K+H2S/A+H2S): Red slant, black in it, yellow butt, bubbles or cracks
  • Alkaline/H2S (K/H2S) : Red slant, black butt
  • Acid/H2S acid: All yellow, little black on bottom
  • Acid/Acid gas (A/A): Yellow slant, yellow butt, air bubbles or cracks
  • Lac operon: any one with yellow slant and yellow butt
34
Q

For Eukaryotes, in what process is RNA polymerase important?

A

Transcription

35
Q

For Eukaryotes, in what process is DNA polymerase important?

A

Replication

36
Q

For Eukaryotes, in what process are ribosomes important?

A

Translation

37
Q

For Eukaryotes, what process occurs in the cytoplasm?

A

Translation

38
Q

For Eukaryotes, what 2 processes occur in the nucleus?

A

Transcription & Replication

39
Q

For Eukaryotes, do chloroplasts and mitochondria replicate their DNA?

A

Yes

40
Q

For Eukaryotes, where does transcription occur?

A

In the mitochondria and the chloroplast

41
Q

For Eukaryotes, where does translation occur?

A

In the mitochondria and the chloroplast

42
Q

How is The structure of Prokaryotic Genomes (genetic information) contained?

A

In two structures;

Chromosomes & Plasmids

43
Q

Are prokaryotes haploid or diploid? Why?

A

Haploid;

because they usually have a single circular chromosome and lack paired homologous chromosomes

44
Q

How is the structure of Eukaryotic Genomes contained?

A

In two structures;

Nuclear DNA & Extranuclear DNA (mitochondria, chloroplast)

45
Q

Which organisms have plasmids and which ones don’t?

A

Fungi, algae, and protozoa have plasmids

Eukaryotes/animals don’t have plasmids

46
Q

What are the 3 general steps that the processes have? (DNA replication, transcription, translation)

A
  1. Initiation
  2. Elongation
  3. Termination
47
Q

DNA replication info:

A
  • Elongation can only occur in the 5’ to 3’ direction

-Continuous strand: on top (leading strand)
-Discontinuous strand: on bottom (lagging strand)

-DNA ligase connects the fragments together

-Linear DNA

48
Q

Bacterial DNA replication info:

A

-Circular DNA

-Has an origin and a termination sequence so it knows when to stop

-Replication forks and proceeds in both directions until the termination

-Breaks off at termination

49
Q

What makes sure that the ends of the linear chromosomes in DNA replication are made?

A

Telomerase

50
Q

Replication info:

Enzyme:
Template:
Start site:
Fidelity mechanism:
Termination:
Location:
Product:
Energy source for process:
Direction of polymerization:

A

Enzyme: DNA polymerases

Template: Both parental strands of DNA

Start site: Origin of replication

Fidelity mechanism: Polymerase proofreading, followed by mismatch repair systems

Termination: Termination sequences

Location: Prokaryotes: cytosol
Eukaryotes: nucleus

Product: 2 daughter DNA strands, each paired with one original strand (semiconservative replication)

Energy source for process: Deoxyribonucleotides

Direction of polymerization: 5’ to 3’

51
Q

Transcription info:

Enzyme:
Template:
Start site:
Fidelity mechanism:
Termination:
Location:
Product:
Energy source for process:
Direction of polymerization:

A

Enzyme: RNA polymerases

Template: 1 strand of DNA

Start site: Promoter

Fidelity mechanism: None

Termination: Terminator

Location: Prokaryotes: cytosol (can be simultaneous w/replication
Eukaryotes: Nucleolus in nucleus

Product: RNA, single stranded

Energy source for process: Ribonucleotides

Direction of polymerization: 5’ to 3’

52
Q

Translation info:

Enzyme:
Template:
Start site:
Fidelity mechanism:
Termination:
Location:
Product:
Energy source for process:
Direction of polymerization:

A

Enzyme: Ribosomes

Template: mRNA

Start site: AUG start codon

Fidelity mechanism: Specificity of enzymes that charge tRNAs

Termination: UAA, UAG, or UGA stop codons

Location: Prokaryotes: cytosol (can be simultaneous w/ transcription

Product: Polypeptides; some function alone as proteins, and others work together as a single protein

Energy source for process: GTP; also ATP for charging tRNAs

Direction of polymerization:
N terminus (end with amino group) to C terminus (end with carboxyl group)

53
Q

Initiation of transcription

A

Prokaryotes:
Consensus sequences in promoters
- Pribnow box (TATAAT): 10bp upstream of start
- 35bp upstream of start (TTGACA)

Bacterial RNA polymerase will recognize the promoter specifically
- Sigma factor helps RNA polymerase find the promoter

Eukaryotes:
Eukaryotes will need transcription factors. These bind first to the promoter then RNA
- TATA box (TATAA): 25-30 bp upstream of start

54
Q

Termination of trancription

A

Prokaryotes:
Stops at the end of the termination signal
- Stem loop structure forms (hits and falls off)
- Rho dependent termination

; Requires the Rho protein which binds to the RNA
; RNA pol pauses at the termination site and Rho causes RNA to be released from DNA

Eukaryotes: Continues for 10-35 nucleotides past the termination signal (AAUAAA)

55
Q

Prokaryotic mRNA

A

(off gene 3)

Organized as one big RNA encoding for many polypeptides (polycistronic)

56
Q

Eukaryotic mRNA

A

One gene = one polypeptide (monocistronic)

  • Starts at: 5’ cap
  • Ends at: 3’ poly A tail
  • Introns have to be cut out
57
Q

What is alternative splicing?

A

When certain introns are retained or some exons are skipped in the final mRNA transcript;

It allows a single gene to produce multiple protein variants or genes by selectively including or excluding specific segments of RNA

58
Q

Initiation

A

Prokaryotes:
- Finds Shine-Dalgarno sequence
- 5’ end of mRNA is complementary to 3’ end of 16S rRNA of ribosome.
This helps it find the Shine-Dalgarno sequence on a polycistronic mRNA

Eukaryotes:
- Finds the 5’ cap
- Downstream of this is AUG, initiating codon, signaling the start of translation

; fMet (prokaryote)
; Met (eukaryote)

59
Q

Elongation

A
  • Amino acids added one at a time
  • Occurs in 3 steps
    ; Codon recognition
    ; Peptide bond formation (23s rRNA ribozyme activity)
    ; Translocation (5’ to 3’ direction)
60
Q

What is the Polyribosome in prokaryotes?

A

In prokaryotes, a polyribosome (or polysome) refers to multiple ribosomes simultaneously translating a single mRNA strand;

While one mRNA molecule is being synthesized, many ribosomes attach to it in succession, each moving along the mRNA to produce its own polypeptide chain.

61
Q

At what levels can gene regulation occur?

A
  • DNA level
  • Transcriptional level
  • Post-transcriptional level
  • Translational level
  • Post-translational level
62
Q

What happens if you don’t do 5’ cap, 3’ poly A tail, and splicing introns?

What happens to the mRNA?

A

The mRNA gets stuck in the nucleus

63
Q

The Operon

Regulatory:
Promoter:
Operator:

A

Its parts

  • Regulatory: gene that encodes for the repressor
  • Promoter: sequence of DNA that tells the RNA polymerase where the beginning of the gene is
  • Operator: sequence of DNA that is between the promoter and the genes of the operon that controls the access of the RNA polymerase to the genes
64
Q

Can RNA molecules control translation?

A

Yes; The Regulatory RNA’s

Examples:

microRNAs (miRNAs): only a portion bind complementary mRNA and inhibit its translation or cleaves mRNA

Small interfering RNA (siRNA): entire molecule complementary to a portion of mRNA, tRNA, or DNA that binds and renders the target inactive by cleaving it

Riboswitch: RNA molecule that changes shape to help regulate translation by blocking initiation or by terminating transcription

Biology 260 (34 decks)

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