BIOL212 W2-2

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Questions and Answers

What is the functional unit of genetic information in microorganisms?

Gene (correct)

Transposable element

Chromosome

Plasmid

What is the primary role of topoisomerases in DNA replication?

To remove supercoils

To synthesize RNA

To introduce supercoils (correct)

To bind to histones

Which statement about plasmids is accurate?

Plasmids do not carry beneficial genes.

Plasmids are found only in eukaryotic cells.

Plasmids are generally linear DNA molecules.

Plasmids replicate independently of chromosomes. (correct)

How is the size of a genome typically expressed?

In base pairs Signup and view all the answers

What percentage of the E. coli K-12 genome is made up of protein-encoding genes?

88% Signup and view all the answers

What type of genetic element usually carries genes for antibiotic resistance?

Plasmids Signup and view all the answers

Which of the following describes the structure of most bacterial chromosomes?

Circular and single Signup and view all the answers

What is the typical gc content of the Escherichia coli K-12 genome?

50.8% Signup and view all the answers

Which element can move from one DNA molecule to another?

Transposable element Signup and view all the answers

What is the effect of supercoiling on DNA structure?

Compacts DNA for accommodation Signup and view all the answers

What is the significance of plasmids in pathogenic bacteria?

Plasmids can carry genes that enhance virulence by producing toxins. Signup and view all the answers

Which statement correctly describes the process of DNA replication?

DNA replication is semiconservative with each double helix containing one parental and one new strand. Signup and view all the answers

What role does DNA polymerase I serve during DNA replication?

It removes RNA primers and replaces them with DNA. Signup and view all the answers

How does the DNA helicase function during DNA replication?

It unwinds the double-stranded DNA ahead of the replication fork. Signup and view all the answers

What is a replisome?

A complex of proteins that facilitates DNA replication. Signup and view all the answers

During DNA replication, how are Okazaki fragments connected?

DNA ligase seals the nicks between the fragments. Signup and view all the answers

What dictates the direction of DNA replication?

Replication occurs from the 5' end to the 3' end of the template strand. Signup and view all the answers

Which of the following statements about bidirectional replication is correct?

It involves two replication forks moving in opposite directions. Signup and view all the answers

What are the precursors for nucleotide synthesis in DNA replication?

Deoxynucleoside 5'-triphosphates Signup and view all the answers

Why do Rhizobia require plasmid-encoded functions?

To fix nitrogen, allowing them to thrive in nutrient-poor environments. Signup and view all the answers

Which components are part of the replisome in DNA replication?

Two copies of DNA polymerase III and helicase Signup and view all the answers

How do DNA polymerases I and III ensure the accuracy of DNA replication?

Through proofreading and 3'→5' exonuclease activity Signup and view all the answers

What is a primary difference between prokaryotic and eukaryotic transcription?

Prokaryotes transcribe multiple genes into a single mRNA Signup and view all the answers

What role does tRNA play in the process of translation?

It carries amino acids to the ribosome to match mRNA codons Signup and view all the answers

What characteristic of mutation rates is generally observed in cells?

Mutation rates range from 10–8 to 10–11 errors per base inserted Signup and view all the answers

During transcription, what must happen to eukaryotic RNAs before translation can occur?

They must be exported from the nucleus Signup and view all the answers

What is the function of primase in DNA replication?

To synthesize a short RNA primer to initiate replication Signup and view all the answers

In what type of cells does coupled transcription and translation occur?

Prokaryotic cells only Signup and view all the answers

Which type of RNA serves as a structural component of the ribosome?

rRNA Signup and view all the answers

What statement accurately describes the process of DNA replication?

Proofreading mechanisms significantly enhance its fidelity Signup and view all the answers

What is the primary function of RNA polymerase in transcription?

To synthesize RNA from a DNA template Signup and view all the answers

Which of the following statements correctly describes bacterial RNA polymerase?

It is a holoenzyme composed of five subunits Signup and view all the answers

What type of RNA does RNA polymerase II primarily transcribe in eukaryotes?

mRNA Signup and view all the answers

Which of the following describes the groove for template DNA binding in bacterial RNA polymerase?

Positively charged to attract negatively charged DNA Signup and view all the answers

What differentiates the various RNA polymerases in eukaryotic organisms?

The specific types of RNA they transcribe Signup and view all the answers

What is a major difference between RNA polymerase and DNA polymerase regarding proofreading?

RNA polymerase does not have a proofreading mechanism Signup and view all the answers

Which RNA polymerase in plants is specifically responsible for generating small interfering RNAs?

Pol IV Signup and view all the answers

Which of the following is required for the activity of RNA polymerase during transcription?

Mg2+ ions Signup and view all the answers

What is generally true about the overall structure of RNA polymerases from eukaryotes and prokaryotes?

They are remarkably similar in structure Signup and view all the answers

What is the role of the positively charged groove in bacterial RNA polymerase?

To aid in the binding of negatively charged DNA Signup and view all the answers

What is the role of the sigma factor in bacterial RNA transcription?

It recognizes and binds to promoter sequences. Signup and view all the answers

Which regions are recognized by the sigma factor in the transcription initiation process?

-10 region and -35 region Signup and view all the answers

In E. coli, what is the significance of the major sigma factor σ70?

It assists in recognizing promoters of related function genes. Signup and view all the answers

How many distinct sigma factors are present in E. coli?

7 Signup and view all the answers

What might indicate a promoter sequence for transcription initiation in bacteria?

A specific length upstream region Signup and view all the answers

What is a characteristic of sigma factors in bacterial species?

The number of sigma factors can vary significantly among species. Signup and view all the answers

What defines the -10 region in RNA transcription initiation?

It is the Pribnow box with about 10 base pairs. Signup and view all the answers

What is the relationship between sigma factors and gene function in bacteria?

Sigma factors enable RNA polymerase to recognize functionally related gene promoters. Signup and view all the answers

Why is the sigma factor described as not tightly bound to the core enzyme?

It facilitates rapid dissociation and reassociation during transcription cycles. Signup and view all the answers

What happens to the sigma factor after transcription initiation has been initiated?

It may dissociate from the core enzyme after initiation. Signup and view all the answers

What is the primary reason that coupled transcription and translation occurs in bacteria but not in eukaryotes?

Eukaryotic transcription occurs in the nucleus, separating it from translation. Signup and view all the answers

What type of termination mechanism relies on an inverted repeat followed by a run of uracils?

Factor-independent termination Signup and view all the answers

What role does the Rho protein play during Rho-dependent termination in bacteria?

It acts as an ATP-dependent RNA/DNA helicase to disengage RNA polymerase. Signup and view all the answers

Which factor is NOT one of the termination factors associated with transcription in E. coli?

Sigma Signup and view all the answers

What happens when Rho protein binds to the rut site during Rho-dependent termination?

It moves along the RNA transcript and disrupts the RNA/DNA hybrid. Signup and view all the answers

Which statement accurately describes a feature of factor-independent termination?

It involves the formation of a hairpin structure that disrupts RNA polymerase binding. Signup and view all the answers

During transcription termination, what occurs when RNA polymerase encounters a transcription terminator sequence?

Transcription halts and RNA polymerase detaches from the template. Signup and view all the answers

What is the primary reason that polymerase pausing can occur at terminator sequences?

The termination sequence is difficult for the polymerase to recognize. Signup and view all the answers

What distinguishes Rho-dependent termination from factor-independent termination?

Rho-dependent termination requires ATP and specific protein interactions. Signup and view all the answers

What triggers the release of the anti-sigma factor NepR, allowing σEcfG1 to function in the transcription process?

Phosphorylation of PhyR Signup and view all the answers

During the formation of the closed complex in transcription, what happens to the DNA?

The DNA remains double-stranded Signup and view all the answers

Which aspect of the sigma factor is crucial for forming the open complex during transcription?

Catalyzing unwinding of the DNA duplex Signup and view all the answers

What describes the composition of a polycistronic mRNA molecule?

Many genes transcribed from a single promoter Signup and view all the answers

What occurs during the elongation phase of transcription?

A transcription bubble of 18 bases forms Signup and view all the answers

In the context of bacterial transcription, what role does the transcription bubble serve?

Keeps RNA polymerase attached to DNA Signup and view all the answers

What characterizes the release of the sigma factor during transcription?

It dissociates after initiation is complete Signup and view all the answers

Which of the following best describes the formation of the open complex during transcription initiation?

Involves localized unwinding at the A/T-rich -10 region Signup and view all the answers

What defines the transcription start site in relation to the first nucleotide of mRNA?

It is defined as position 0 or +1 Signup and view all the answers

Which factor is crucial for transcription initiation in bacterial cells?

Sigma factors Signup and view all the answers

What best describes the process that allows proteins to begin to form before mRNA is fully transcribed in bacteria?

Coupled transcription and translation Signup and view all the answers

What outcome typically occurs when RNA polymerase encounters a transcription terminator?

Transcription terminates Signup and view all the answers

In factor-independent termination, which sequence structure is commonly formed?

A hairpin structure Signup and view all the answers

Which statement is true about Rho-dependent termination in E. coli?

It relies on Rho binding to rut sites Signup and view all the answers

What role does the Rho hexamer play in transcription termination?

It acts as an ATP-dependent RNA helicase Signup and view all the answers

What consequence might arise from incorrect recognition of terminator sequences?

The RNA polymerase continues transcription indefinitely Signup and view all the answers

Why do eukaryotes not exhibit coupled transcription and translation similar to bacteria?

Transcription occurs in the nucleus and translation in the cytoplasm Signup and view all the answers

Which of the following statements about transcription termination in bacteria is correct?

Both intrinsic and Rho-dependent methods exist Signup and view all the answers

What is the function of the rut site in the context of Rho-dependent termination?

It acts as a recognition site for Rho Signup and view all the answers

What characterizes the distinction between prokaryotic and eukaryotic transcription processes?

Eukaryotic transcription occurs in the nucleus Signup and view all the answers

What initiates the release of the anti-sigma factor NepR in M.extorquens during stress response?

Phosphorylation of PhyR Signup and view all the answers

What occurs during the formation of the open complex in transcription initiation?

The DNA unwinds in the A/T-rich -10 region Signup and view all the answers

Which statement best describes the elongation phase of transcription?

An RNA/DNA hybrid is formed during RNA synthesis Signup and view all the answers

What is a characteristic feature of polycistronic mRNA?

It encodes multiple genes with related functions from a single promoter Signup and view all the answers

Which factor plays a crucial role in preventing the need for a helicase during transcription initiation?

Sigma factor's aromatic residues Signup and view all the answers

In which scenario can transcription occur simultaneously on the same gene?

Only if multiple RNA polymerases are recruited to the promoter region Signup and view all the answers

Which feature distinguishes the initiation of transcription at the +1 site?

The RNA transcript starts with a triphosphate group Signup and view all the answers

What role do anti-sigma factors serve in cellular processes?

They inhibit sigma factors to regulate gene expression Signup and view all the answers

What dictates the directionality of transcription during gene expression?

The orientation of the gene within the DNA double helix Signup and view all the answers

The transient melting of the DNA duplex during transcription is primarily facilitated by which component?

Aromatic residues in the sigma factor Signup and view all the answers

Study Notes

DNA and Genetic Information Flow

The functional unit of genetic information is the gene.
Genes are part of genetic elements, which are large molecules, such as chromosomes.
Nucleic acids contain genetic information through nucleotides, which are monomers of nucleic acids.
DNA is the genetic blueprint, and RNA is its transcription product.
Both DNA and RNA are polynucleotides.
Informational macromolecules include nucleic acids and proteins.

Genetic Elements: Chromosomes and Plasmids

Genome size is expressed in base pairs.
- 1000 base pairs = 1 kilobase pair = 1 kbp
- 1 million base pairs = 1 megabase pair = 1 Mbp
The E. coli genome is 4.64 Mbp and encodes around 4000 genes.
Linear DNA is several hundred times longer than the cell, so supercoiling compacts DNA to accommodate the genome.
Topoisomerases insert and remove supercoils.
DNA gyrase introduces supercoils into DNA via double-strand breaks.
Chromosomes are the main genetic element in prokaryotes.
Most bacteria and archaea have a single circular chromosome carrying all or most of their genes.
Plasmids are double-stranded DNA that replicate separately from the chromosome.
- Usually circular.
- Generally beneficial for the cell (e.g., antibiotic resistance).
- Not extracellular, unlike viruses.
Transposable elements are segments of DNA that can move from one site to another on the same or a different DNA molecule.
- They can be inserted into other DNA molecules (e.g., chromosomes, plasmids, viral genomes).

Escherichia coli K-12 Chromosome Features

Approximately 5 Mbp in size.
GC content (% of G and C in the genome) is 50.8 %.
Almost 4300 possible protein-encoding genes, comprising 88% of the genome.

Plasmid Features

Plasmids can carry genes associated with virulence factors in pathogenic bacteria that contribute to infection.
Bacteriocins, proteins that inhibit or kill closely related species or different strains of the same species, can be encoded on plasmids.
Rhizobia use plasmid-encoded functions to fix nitrogen.
Plasmids can contribute to metabolism, such as hydrocarbon degradation.
Plasmids are crucial for conjugation, a form of horizontal gene transfer.

### DNA Replication

DNA replication is semiconservative, meaning each new double helix consists of one original strand and one newly synthesized strand.
Deoxynucleoside 5′-triphosphate (dNTP) serves as the precursor for each nucleotide.
DNA replication always proceeds from the 5′ end to the 3′ end.
DNA polymerases, specifically DNA Pol III, are the primary enzymes responsible for replicating chromosomal DNA in E. coli.
Other DNA polymerases play roles in DNA repair.
DNA synthesis initiates at the origin of replication in prokaryotes.
DNA polymerases require a primer, a short stretch of RNA, which primase creates.
The replication fork is the region of unwound DNA where replication occurs.
DNA helicase unwinds the DNA.
DNA synthesis occurs continuously on the leading strand and discontinuously on the lagging strand.
Okazaki fragments are short segments of DNA synthesized on the lagging strand.
DNA polymerase I removes the RNA primer and replaces it with DNA.
DNA ligase seals the gaps in the DNA.
Bidirectional replication occurs in circular chromosomes.
Two replication forks move in opposite directions along the circular chromosome.
DNA Pol III adds approximately 1,000 nucleotides per second.
The replisome, a complex of multiple proteins involved in replication, consists of two copies of DNA polymerase III, DNA gyrase, helicase, primase, and multiple single-strand DNA-binding proteins.
DNA replication is extremely accurate due to proofreading mechanisms that ensure high fidelity.
Mutation rates in cells are typically 10–8 to 10–11 errors per base inserted.
DNA Pol I and Pol III can detect mismatches through incorrect hydrogen bonding and remove them via their 3'→5' exonuclease activity.

RNA Synthesis: Transcription

Central Dogma: DNA is duplicated (replication), information is transferred from DNA to RNA (transcription), and information in RNA is used to build polypeptides (translation).
mRNA (messenger RNA) encodes proteins.
tRNA (transfer RNA) converts mRNA into the amino acid sequence of a protein.
rRNA (ribosomal RNA) provides catalytic and structural components for ribosomes.

### Transcription Characteristics in Prokaryotes & Eukaryotes

Eukaryotes: Each gene is transcribed individually into a single mRNA. Replication and transcription happen in the nucleus. RNA must be exported outside the nucleus for translation.
Prokaryotes: Multiple genes can be transcribed in a single mRNA. Transcription and translation are coupled, maximizing protein synthesis.

Transcription in Bacteria

Transcription is the synthesis of RNA from a DNA template.
RNA polymerase is the enzyme that carries out transcription.
RNA polymerase requires DNA as a template and uses ATP, GTP, CTP, and UTP as RNA precursors.
Transcription proceeds in the 5' to 3' direction, similar to DNA replication.
Only one strand of DNA is transcribed.
No primer is required for RNA transcription.

Prokaryotic vs. Eukaryotic RNA Polymerases

Most eukaryotes have three different RNA polymerases:
- Pol I: synthesizes ribosomal RNAs (rRNAs)
- Pol II: synthesizes messenger RNAs (mRNAs)
- Pol III: synthesizes 5S rRNA, transfer RNAs (tRNAs), and other small RNAs
Plants have two additional RNA polymerases:
- Pol IV: involved in the generation of small interfering RNAs (siRNAs) from transposons and repetitive sequences, contributing to epigenetic regulation.
- Pol V: responsible for silencing (DNA and histone methylation) of transposons and repetitive sequences.
Bacteria and Archaea only have a single RNA polymerase.

Bacterial RNA Polymerase Structure

E. coli RNA polymerase is composed of five subunits: two α (αI and αII), β, β', and ω.
The core enzyme consists of the β, β', and ω subunits.
Together, the core enzyme and the sigma factor form the holoenzyme.

Key Features of Bacterial RNA Polymerase

The DNA binding groove of RNA polymerase is positively charged, while DNA is negatively charged, facilitating the binding of DNA.
Magnesium ions (Mg2+) are required for RNA polymerase activity.
RNA polymerase lacks a proofreading mechanism, unlike DNA polymerase, which has 3' to 5' exonuclease activity.

Transcription Initiation in Bacteria

Initiation is sequence-specific and occurs at the promoter sequence, located upstream of the RNA transcription start site.
The core enzyme can catalyze transcription, but the sigma factor is needed for promoter recognition.
Sigma factors are bacterial transcription factors that bind to the promoter sequence (-10 and -35 regions) and help RNA polymerase initiate transcription.
The +1 site marks the transcription initiation site.
The -10 region (also known as the Pribnow box) is a consensus sequence located about 10 base pairs upstream of the +1 site.
The -35 region is another consensus sequence located about 35 base pairs upstream of the +1 site.
Multiple sigma factors exist in bacteria, allowing RNA polymerase to recognize and transcribe different sets of genes.

Sigma Factor in E. coli

E. coli has seven distinct sigma factors.
The primary sigma factor, σ70, is responsible for the transcription of "housekeeping" genes.
Each sigma factor typically binds to a set of genes with related functions, such as sporulation genes.

Examples of Sigma Factor Functions

Sporulation sigma factor in Bacillus subtilis: responsible for transcribing genes involved in bacterial sporulation in response to adverse conditions.

RNA Polymerase Holoenzyme Structure

The sigma factor is attached to the core RNA polymerase complex.

Anti-sigma Factors

Inhibit sigma factor function by binding to them
Regulate stress response in M.extorquens
Phosphorylation of PhyR releases anti-sigma factor (NepR) from σEcfG1
Allows σEcfG1 to associate with RNA polymerase and transcribe stress genes

Transcription Start and Elongation

Closed Complex: DNA is double-stranded, RNA polymerase is bound to the promoter
Open Complex: Sigma factor unwinds DNA at the promoter region, no helicase or ATP required
Initiating Ribonucleotides: Transcription usually starts at +1 with a nucleotide triphosphate, resulting in a triphosphate at the 5' end of the newly synthesized RNA
Elongation:
- Sigma subunit detaches from the polymerase
- Transcription bubble forms (18 base long DNA opening)
- RNA/DNA hybrid forms, helping keep the polymerase attached to the DNA

Transcription Features

Transcription can occur on either DNA strand
Multiple RNA polymerases can transcribe the same gene simultaneously

Polycistronic mRNA

Single mRNA molecule containing multiple genes
Transcribed from the same promoter upstream of a cluster of genes with related functions (operon)

Ribosomal RNA

Polycistronic

Coupled Transcription and Translation in Bacteria

Ribosomes attach to the 5' end of mRNA before transcription is complete
Ribosomes move along the mRNA towards the 3' end as transcription continues
Allows protein synthesis to begin before mRNA is fully transcribed
Not possible in eukaryotes due to separate compartments for transcription and translation

Transcription Termination

RNA polymerase detaches from the template when it encounters a terminator sequence
Two types: factor-independent and Rho-dependent

Factor-Independent Termination

Inverted repeat sequence with a central non-repeating segment forms a hairpin structure
Followed by a run of U nucleotides
Destabilizes RNA/DNA binding, leading to RNA dissociation

Rho-Dependent Termination

Requires Rho factor - an RNA-dependent ATPase and RNA/DNA helicase
Recognizes rut sites in RNA
Rho binds to rut and acts as a helicase, moving 5' to 3' along the transcript
This pulls apart the RNA/DNA hybrid, releasing the RNA polymerase when it's paused at the terminator sequence