Unit 2 Bio Flashcards
(218 cards)
1
Q
What is replication?
A
Process of taking one DNA molecule and producing an identical copy of it.
2
Q
When does replication occur
A
Replication occurs during the S-phase of the cell cycle,
when a cell produces identical copies of all of its chromosomes.
3
Q
Who discovered that chromosomes appeared to carry genes.
A
TH Morgan
4
Q
What did F Griffith discover?
A
He found that giving mice a mixture of heat-
treated S-type (lethal bacterial) and untreated R-type (non lethal bacteria) would kill mice. This
discovery indicated that genes could be transferred between organisms.
5
Q
Who were the first to determined that DNA was the molecule responsible for the transformation of bacteria.
A
Oswald Avery, Maclyn McCarty and Colin MacLeod
6
Q
Who proved with bacteriophages that DNA was responsible for the encodement of genes.
A
Alfred Hershey and Martha Chase
7
Q
What was the experiment done by Alfred Hershey and Martha Chase.
A
They used bacteriophages with either 35 S-radioactive isotope of sulphur in their proteins or 32 P-radioactive isotope in DNA and measured which was present in infected bacteria. It was found to be the phosphorus.
8
Q
What is conservative replication?
A
Where the initial molecule remains intact, and the second molecule is a copy consisting of entirely new DNA
9
Q
What is semi-conservative replication?
A
Where the initial molecule separates and each half is used to form a new strand, resulting in two molecules that each contain one original strand and one “new” strand.
10
Q
What is dispersive replication
A
Where fragments of the initial dsDNA are incorporated throughout the two dsDNA molecules produced so that after every round of replication, each molecule contains a fraction of the original dsDNA molecule
11
Q
What is the Meselson and Stahl experiment?
A
They produced DNA in an environment with 15N and then moved it to an environment with 14N. Then DNA was produced. The DNA were separated by centrifugation, to see if they had different weights. After the first round of replication, both dsDNA molecules produced had the same weight, producing only a single band. A second round was performed and two bands were formed. Proving semi-conservative.
12
Q
Which strand of DNA is copied continuously?
A
3’ strand or leading strand
13
Q
What is the role of DNA Helicase in DNA replication?
A
DNA Helicase unwinds the double helix by breaking the hydrogen bonds between complementary base pairs
14
Q
Why are single-stranded binding proteins (SSBPs) important during replication?
A
SSBPs bind to single-stranded DNA to prevent it from reannealing or forming secondary structures.
15
Q
How does Topoisomerase prevent coiled tension during DNA replication?
A
Topoisomerase relieves the supercoiling tension caused by helicase by cutting, untwisting, and rejoining the DNA strands.
16
Q
What is the purpose of Primase in DNA replication?
A
Primase synthesizes a short RNA primer that provides a free 3’ hydroxyl group for DNA Polymerase III to begin DNA synthesis.
17
Q
Why can DNA Polymerase III not bind directly to single-stranded DNA?
A
DNA Polymerase III requires an RNA primer to provide a free 3’ hydroxyl group for nucleotide addition.
18
Q
In what direction does DNA Polymerase III add nucleotides?
A
DNA Polymerase III adds nucleotides in the 5’ to 3’ direction.
19
Q
What are Okazaki fragments, and why are they formed?
A
Okazaki fragments are short DNA sequences on the lagging strand, formed because DNA Polymerase III can only synthesize DNA in the 5’ to 3’ direction.
20
Q
Why is DNA replication on the lagging strand described as discontinuous?
A
Since replication must occur in the 5’ to 3’ direction, the lagging strand is synthesized in short, separate Okazaki fragments.
21
Q
What enzyme removes RNA primers and replaces them with DNA nucleotides?
A
DNA Polymerase I removes RNA primers and replaces them with DNA nucleotides.
22
Q
What enzyme removes RNA primers and replaces them with DNA nucleotides?
A
DNA Polymerase I removes RNA primers and replaces them with DNA nucleotides.
23
Q
How does DNA Ligase contribute to the completion of DNA replication?
A
DNA Ligase seals the gaps between Okazaki fragments by forming phosphodiester bonds.
24
Q
Why is the lagging strand replicated more slowly than the leading strand?
A
The lagging strand is synthesized discontinuously, requiring more frequent primer synthesis, fragment formation, and ligation.
25
How does DNA Polymerase III reduce errors during replication?
DNA Polymerases actually have proof-reading capabilities as they are forming phosphodiester bonds. This means that if DNA Pol recognizes that it has base-paired the wrong nucleotide, that faulty nucleotide will be removed and the correct partner will instead be added.
26
What is the role of DNA Repair Machinery?
DNA Repair Machinery detects and fixes DNA damage by excising incorrect bases and replacing them with the correct ones.
27
Explain how a nuclease is involved in DNA repair.
A nuclease cuts and removes damaged or mismatched DNA segments during nucleotide excision repair.
28
After the damaged DNA is removed, how are the correct nucleotides added back?
DNA Polymerase I adds the correct nucleotides by using the undamaged strand as a template.
29
Why is DNA Ligase needed in the DNA repair process?
DNA Ligase seals the repaired DNA by forming phosphodiester bonds at the nicks.
30
What is Xeroderma Pigmentosum, and how does it relate to DNA repair deficiencies?
Xeroderma Pigmentosum is a genetic disorder caused by defective nucleotide excision repair enzymes, leading to extreme sensitivity to UV light and a high risk of skin cancer.
31
How do single-strand DNA breaks differ from double-strand breaks in terms of repair mechanisms?
Single-strand breaks are repaired using the complementary strand as a template, while double-strand breaks may be repaired using homologous recombination or non-homologous end joining.
32
Why might a homologous chromosome be used for repair in some cases?
During homologous recombination, a homologous chromosome serves as a template to accurately repair a double-strand break.
33
What are Restriction Endonucleases?
Enzymes capable of hydrolyzing phosphodiester bonds in dsDNA at very specific sequences called “restriction sites”.
34
How do bacteria protect themselves from endonucleases used to attack viruses?
Bacteria either avoid having the specific nucleotide sequence recognized by
the restriction endonucleases, or they methylate their DNA, so that it cannot be recognized by the enzymes.
35
Do single-celled organisms commit aptoptosis?
No, they tend to rely on a process known as the SOS response, whereby drastic solutions are attempted to repair damaged DNA.
36
Do prokaryotes possess telomeres?
No, they do not as they do not have "ends" to their chromosomes. Their circular DNA prevents them from having such segments.
37
Why do telomeres get shorter?
On the lagging strand, an RNA primer will exist at the very end, but this will be removed during replication. Unfortunately, since no dsDNA precedes this region, DNA Pol I cannot properly bind to add DNA nucleotides to replace the RNA
primer that is removed
38
What are telomeres?
Repeating units of non-coding DNA at the two ends of each chromosome, called telomeres. Telomeres consist of a sequence of TTAGGG in repeats of 100 to over 1000 copies per chromosome.
39
How do telomeres limit the rounds of replication possible for a cell?
Once telomeres "lose" enough length, the cell does not replicate anymore, as its genetic material would be compromised. Therefore, cell aging is linked to telomere length.
40
What is the solution for cells to prevent telomere shortening?
Telomerase
41
What is telomerase and how does it function?
Telomerase is an enzyme that allows telomeres to be extended. Telomerase contains a protein and nucleic acid component. The nucleic acid portion contains a sequence of nucleotides that are complementary to telomeres. This sequence allows Telomerase to bind to a shortening telomere, and Telomerase then catalyses the addition of nucleotides to elongate the telomere.
42
What are the cells that possess telomerase?
Reproductive cells, white blood cells
and stem cells.
43
What is Werner Syndrome, and how does it relate to telomeres?
Werner Syndrome is a condition where individuals cannot maintain their telomeres properly, leading to premature aging.
44
How might telomerase be related to cancer?
Many cancer cells express telomerase to overcome the normal limits on cell division.
45
What is chromatin?
The complex of DNA and proteins (histones) that form chromosomes.
46
What are nucleosomes?
DNA segments wrapped around histone octamers, forming the basic unit of chromatin.
47
What is the difference between euchromatin and heterochromatin?
Euchromatin is loosely packed and transcriptionally active, while heterochromatin is tightly packed and transcriptionally inactive.
48
How does histone acetylation affect chromatin structure?
It reduces the positive charge on histones, decreasing their interaction with DNA and promoting a more open, euchromatic state.
49
What enzymes add acetyl groups to histones?
Histone Acetyl Transferases (HATs).
50
Which enzymes remove acetyl groups from histones?
Histone Deacetylases (HDACs).
51
How does histone methylation generally affect chromatin?
It usually increases chromatin packing, making it less accessible for transcription.
52
What is the effect of histone phosphorylation on chromatin structure?
It promotes the formation of euchromatin by introducing negative charges that repel DNA.
53
How can DNA methylation regulate gene expression?
Methylation of cytosine residues in DNA generally silences gene expression by blocking transcription factor binding.
54
What is epigenetics?
The study of heritable changes in gene function that do not involve changes in the DNA sequence, often through DNA and histone modifications.
55
How does the epigenome affect cellular differentiation?
It determines which genes are expressed or silenced, guiding a cell’s development into a specific type
56
What is the function of RNA Polymerase II?
It synthesizes messenger RNA (mRNA) from a DNA template during transcription.
57
What is the TATA box?
A conserved DNA sequence (TATAAAA) in the promoter region where RNA Polymerase II and transcription factors bind to initiate transcription.
58
What role do transcription factors play in gene expression?
They bind to specific DNA sequences to either activate or repress transcription.
59
What are enhancers?
Distal control elements that, when bound by activator proteins, increase transcription levels.
60
How does the repressor protein function in the Trp operon?
When activated by tryptophan, it binds to the operator and blocks transcription of tryptophan synthesis genes.
61
What is the default state of a repressible operon?
The default is “on” (transcription occurs) unless the repressor binds the operator.
62
How does the Lac operon illustrate inducible regulation?
The Lac repressor normally binds the operator and blocks transcription; in the presence of lactose, the repressor is inactivated, allowing transcription
63
Which three genes are part of the Lac operon?
lacZ, lacY, and lacA.
64
What enzyme does lacZ encode and what is its function?
β-galactosidase, which hydrolyzes lactose into glucose and galactose.
65
What is the role of the lacY gene product?
Permease, which transports lactose into the bacterial cell.
66
What is the lacA gene product, and what is known about its function?
Transacetylase; its exact function is not clearly understood.
67
How does the Catabolic Activator Protein (CAP) regulate the Lac operon?
CAP binds to RNA polymerase to stabilize its attachment to the DNA; its activation is dependent on cAMP levels.
68
What does an increase in cAMP indicate about a bacterial cell’s energy state?
High cAMP levels indicate low glucose, promoting CAP activation and the transcription of the Lac operon (if lactose is present).
69
What is the overall advantage of operon-based regulation in prokaryotes?
It allows for the coordinated and efficient regulation of multiple genes that function together in a metabolic pathway.
70
How does the regulation of transcription differ between prokaryotes and eukaryotes?
Prokaryotes often use operons with simple on/off mechanisms, while eukaryotes have complex regulation involving chromatin remodeling, multiple transcription factors, and epigenetic modifications.
71
What is the role of the 5’ cap in eukaryotic mRNA?
It protects mRNA from degradation, assists in nuclear export, and helps in ribosome recruitment for translation.
72
How is the 5’ cap added to mRNA?
Shortly after transcription begins, a modified guanine nucleotide (methyl G cap) is added to the 5’ end.
73
What is the purpose of the 3’ polyA tail in mRNA?
It protects mRNA from degradation, aids in nuclear export, and promotes translation initiation.
74
How is the polyA tail added to mRNA?
After transcribing the polyadenylation signal (AAUAAA), specific proteins cleave the mRNA and add a series of adenine nucleotides.
75
What is splicing and why is it important in eukaryotes?
Splicing removes introns from the pre-mRNA to produce a mature mRNA that can be translated into protein.
76
What are introns and exons?
Introns are non-coding regions removed during splicing; exons are coding sequences that remain in the mature mRNA.
77
What complex carries out the splicing of pre-mRNA?
The spliceosome.
78
What are snRNAs and what is their role in splicing?
Small nuclear RNAs (snRNAs) are part of the spliceosome; they help recognize splice sites by base pairing with the pre-mRNA.
79
80
What is alternative splicing?
A process where the splicing pattern varies to produce different mature mRNAs from the same gene, increasing protein diversity.
81
Why is alternative splicing important for gene expression?
It allows one gene to encode multiple protein variants with different functions.
82
What is mRNA localization?
The process by which mRNAs are directed to specific regions within the cytoplasm for localized translation.
83
How do the untranslated regions (UTRs) of mRNA affect its fate?
UTRs contain signals that regulate mRNA stability, localization, translation efficiency, and degradation.
84
What is the role of RNA Polymerase II in transcription?
It synthesizes mRNA from the DNA template.
85
What are the three main stages of transcription?
Initiation, elongation, and termination.
86
During transcription initiation, where is RNA Polymerase II recruited?
To the promoter region, specifically upstream at the TATA box.
87
How does RNA Polymerase II separate the DNA strands?
It unwinds a short region of the DNA to expose the template strand for transcription.
88
In which direction does RNA Polymerase II move along the DNA template during elongation?
It moves in the 3’ to 5’ direction, synthesizing RNA in the 5’ to 3’ direction
89
What signals termination in eukaryotic transcription?
The polyadenylation signal (AAUAAA) followed by a short run of additional nucleotides.
90
How is transcription termination achieved in prokaryotes?
By specific terminator sequences that signal RNA Polymerase to stop transcription.
91
What are the roles of RNA Polymerase I and III?
RNA Polymerase I synthesizes rRNA and RNA Polymerase III produces tRNA and other small RNAs.
92
What is a transcription factor?
A protein that binds specific DNA sequences to regulate the rate of transcription of genetic information.
93
What is the DNA-binding domain of a transcription factor?
The region that enables the transcription factor to bind to specific DNA sequences.
94
What is the activation domain of a transcription factor?
The region that interacts with other proteins to modulate transcription, such as recruiting or blocking RNA Polymerase.
95
How can transcription factors influence chromatin structure?
By recruiting chromatin modifiers like HATs or HDACs, thereby altering nucleosome spacing and DNA accessibility.
96
What is feedback regulation in gene expression?
A mechanism where the end product of a pathway regulates its own synthesis by influencing transcription initiation.
97
What is an operon?
A cluster of genes under the control of a single promoter and operator, common in prokaryotes.
98
How does the Trp operon function?
When tryptophan is abundant, it binds the repressor protein which then binds the operator to shut off gene transcription.
99
What happens to the Trp operon when tryptophan levels are low?
The repressor remains inactive, allowing RNA Polymerase to transcribe the tryptophan synthesis genes.
100
How is the default state of the Lac operon described?
The repressor is active by default, blocking transcription unless inactivated by lactose.
101
How does lactose affect the Lac operon?
Lactose binds to the repressor, inactivating it and allowing transcription of genes for lactose metabolism.
102
What role does CAP play in the Lac operon regulation?
CAP binds to the promoter region to facilitate RNA Polymerase binding when glucose levels are low.
103
Why is glucose level important in regulating the Lac operon?
Low glucose increases cAMP, which activates CAP; high glucose results in low cAMP, reducing CAP activation.
104
What is the significance of having both activators and repressors in transcriptional regulation?
They provide fine-tuned, condition-dependent control over gene expression.
105
What are the key differences between polycistronic and monocistronic mRNA?
Prokaryotic mRNAs are polycistronic (multiple coding regions per transcript), whereas eukaryotic mRNAs are typically monocistronic (one coding region per transcript).
106
What is co-transcriptional translation?
In prokaryotes, translation begins before transcription is completed, with ribosomes binding to the mRNA as it is synthesized.
107
What role does the cap-binding complex play in mRNA export?
It recognizes the 5’ cap and facilitates the transport of mRNA through the nuclear pore.
108
How does the polyA tail contribute to mRNA stability?
It protects the mRNA from degradation by exonucleases and assists in translation initiation.
109
What enzyme cleaves the mRNA downstream of the polyadenylation signal?
A specific endonuclease associated with the polyadenylation machinery.
110
What is the typical length of a polyA tail in eukaryotic mRNA?
It can range from 50 to over 250 nucleotides.
111
What is the function of the 5’ untranslated region (UTR)?
It regulates translation initiation and mRNA stability without being translated into protein.
112
What is the role of the 3’ untranslated region (UTR) in mRNA?
It contains regulatory sequences that affect mRNA localization, stability, and translation efficiency.
113
What structural feature do tRNAs possess that is crucial for translation?
The anticodon loop, which base pairs with codons on the mRNA.
114
What is the function of the amino acid attachment site on tRNA?
It is where the corresponding amino acid is covalently linked by aminoacyl tRNA synthetases.
115
How many different aminoacyl tRNA synthetases are there and why?
There are 20, one for each amino acid.
116
What is the “wobble” hypothesis in translation?
It explains how some tRNAs can recognize more than one codon due to flexible base pairing at the third codon position.
117
What is the structure and function of the ribosome?
A ribonucleoprotein complex that facilitates the translation of mRNA into protein; it has two subunits (small and large) and three tRNA binding sites (A, P, and E).
118
What is the function of the A-site in the ribosome?
It is the entry site for aminoacyl-tRNAs carrying the next amino acid.
119
What is the role of the P-site in the ribosome?
It holds the tRNA carrying the growing polypeptide chain.
120
What happens at the E-site of the ribosome?
It is the exit site for tRNAs that have released their amino acids.
121
What initiates translation in eukaryotes?
The binding of the methionine-charged initiator tRNA to the 40S ribosomal subunit at the start codon (AUG).
122
What energy molecule is used during the initiation of translation?
GTP.
123
Describe the process of elongation during translation.
Aminoacyl-tRNAs enter the A-site, peptide bonds form between the amino acid in the P-site and the new amino acid, then the ribosome translocates, moving tRNAs from the A-site to the P-site and ejecting the empty tRNA from the E-site.
124
What catalyzes peptide bond formation in the ribosome?
The rRNA component of the ribosome acts (petptidyl transferase) as a ribozyme to catalyze peptide bond formation.
125
What signals translation termination?
Stop codons, which are recognized by release factors rather than tRNAs.
126
What role do release factors play in translation termination?
They bind to stop codons and trigger the release of the completed polypeptide chain, as well as the disassembly of the ribosome.
127
What is a polysome?
A complex formed by multiple ribosomes simultaneously translating a single mRNA molecule.
128
How does mRNA circularization enhance translation efficiency?
Circularization brings the 5’ cap and 3’ polyA tail into close proximity, facilitating ribosome recycling and rapid reinitiation.
129
How is general translation inhibited under cellular stress?
Through the inactivation of the initiation factor eIF2, which reduces the initiation of new protein synthesis.
130
What are the two main locations for translation within a eukaryotic cell?
In the cytosol (free ribosomes) and on the rough endoplasmic reticulum.
131
How does mRNA degradation help regulate gene expression?
By controlling the lifespan of mRNA molecules, thus determining how long they are available for translation.
132
What triggers the degradation of an mRNA molecule?
The removal of protective structures (5’ cap or polyA tail) which then allows exonucleases to degrade the mRNA.
133
Which enzymes remove the 5’ cap and the polyA tail during mRNA degradation?
Decapping enzymes remove the 5’ cap and deadenylases remove the polyA tail.
134
What is the role of exonucleases in mRNA degradation?
They degrade the mRNA from the exposed ends once the cap or tail has been removed.
135
What is the significance of microRNAs (miRNAs) in mRNA regulation?
miRNAs bind to complementary sequences on mRNAs to direct their degradation or inhibit their translation
136
What is the relationship between mRNA stability and protein production?
More stable mRNAs tend to produce more protein over time, while rapidly degraded mRNAs result in lower protein levels.
137
How does the concept of “degeneracy” apply to the genetic code?
Multiple codons can code for the same amino acid.
138
How many possible codons exist and how many amino acids do they encode?
There are 64 possible codons for 20 amino acids (plus stop codons).
139
Why is the genetic code considered nearly universal?
Because nearly all organisms use the same codon-to-amino acid mapping.
140
What is the functional significance of the anticodon in tRNA?
It ensures that the correct amino acid is added by base pairing with the corresponding codon on the mRNA.
141
What is the importance of the aminoacyl-tRNA synthetase enzymes’ specificity?
They ensure that each tRNA is charged with the correct amino acid, maintaining the fidelity of translation.
142
How do aminoacyl-tRNA synthetases “activate” an amino acid?
By using ATP to form an aminoacyl-AMP intermediate before transferring the amino acid to the tRNA.
143
What is “wobble base pairing” and why is it important?
It allows some tRNAs to recognize multiple codons for the same amino acid, reducing the number of distinct tRNA species needed.
144
What structural feature of tRNA allows it to fold into its characteristic cloverleaf shape?
Its specific nucleotide sequence and intramolecular base pairing interactions.
145
How does the structure of the ribosome facilitate its function in translation?
Its two subunits create distinct sites (A, P, E) that coordinate tRNA binding, peptide bond formation, and translocation.
146
What is the role of rRNA within the ribosome?
rRNA not only provides structural support but also catalyzes the formation of peptide bonds.
147
How does the ribosome ensure reading frame fidelity during translation?
Through precise interactions between codons on the mRNA and anticodons on tRNAs, along with ribosomal proofreading mechanisms.
148
What determines which mRNAs are translated at the rough ER versus in the cytosol?
Signal sequences in the mRNA or nascent protein direct the ribosome to the endoplasmic reticulum.
149
What is the significance of coupling between transcription and RNA processing in eukaryotes?
It ensures that only properly processed mRNAs (with a cap, tail, and spliced exons) are exported and translated.
150
Why is it important that splicing occurs before mRNA export in eukaryotes?
It prevents the translation of incomplete or faulty mRNA transcripts.
151
How do cells ensure that methylation patterns are maintained after DNA replication?
DNA Methyl Transferases (DNMTs) add methyl groups to the newly synthesized strand to mirror the parent strand.
152
What is the effect of global DNA methylation on gene expression?
It generally leads to long-term gene silencing.
153
How can changes in histone modifications affect memory formation and learning?
Epigenetic modifications such as histone acetylation can influence gene expression patterns that are critical for synaptic plasticity.
154
How does chromatin remodeling affect transcription factor access to DNA?
Loosening chromatin (euchromatin) increases accessibility, while tight packing (heterochromatin) restricts access.
155
What is the role of control elements in the regulation of transcription?
They bind transcription factors that enhance or repress transcription from the promoter.
156
How do proximal control elements differ from distal control elements?
Proximal elements are near the promoter, while distal elements (enhancers) can be thousands of nucleotides away yet still affect transcription.
157
How can transcription factors alter the orientation or folding of DNA?
They can induce bends or loops in the DNA, bringing distant regulatory elements into close proximity with the promoter.
158
What is the significance of the nuclear pore in mRNA export?
It regulates which mRNAs (properly processed and capped/tail-equipped) are allowed to exit the nucleus.
159
How do proteins associated with the 3’ UTR influence mRNA fate?
They can either promote translation by recruiting initiation factors or signal for degradation by recruiting decapping/deadenylation enzymes.
160
What role does the spliceosome play in ensuring accurate mRNA splicing?
It recognizes conserved splice sites at exon–intron boundaries and precisely excises introns while joining exons.
161
How can errors in splicing affect protein function?
Mis-splicing can lead to frameshifts or the exclusion/inclusion of critical exons, altering the resulting protein’s structure and function.
162
How does the nuclear pore complex contribute to mRNA quality control?
It selectively permits only properly processed mRNAs to exit by assessing the presence of a 5’ cap and polyA tail and by requiring binding to specific transport proteins.
163
What is the significance of mRNA localization within the cytosol?
Localization allows mRNAs to be directed to specific cellular regions for localized translation, which is essential for processes like cell movement.
164
What roles do the 5’ and 3’ untranslated regions (UTRs) play in mRNA regulation?
UTRs act as “docking sites” for proteins that regulate mRNA localization, translation initiation, and degradation.
165
How is the correct amino acid attached to its tRNA?
Aminoacyl-tRNA synthetases catalyze the attachment of the correct amino acid to its corresponding tRNA using ATP.
166
What is a polysome, and why is it important?
A polysome is a cluster of ribosomes translating the same mRNA simultaneously, which increases the efficiency of protein synthesis.
167
How does mRNA circularization benefit translation efficiency?
Circularization brings the 5’ cap and 3’ polyA tail into close proximity, facilitating ribosome recycling and rapid reinitiation of translation.
168
What role does eIF2 play in translation initiation?
eIF2 is a key initiation factor that helps recruit the initiator tRNA to the ribosome; its inhibition during stress reduces overall protein synthesis.
169
Where within the cell are ribosomes typically found, and how does this relate to protein targeting?
Ribosomes are found free in the cytosol or bound to the rough endoplasmic reticulum, which directs proteins to different cellular destinations.
170
What are the two major phases of protein processing in cells?
Protein processing involves proper folding/localization and post-translational modifications, which together dictate a protein’s function, stability, and cellular destination.
171
How do signal sequences contribute to protein localization?
A signal sequence is a short peptide at the beginning of a protein that directs the ribosome to the endoplasmic reticulum (ER) for proteins destined for secretion or membrane insertion; it is later cleaved off.
172
What role does the Signal Recognition Particle (SRP) play in protein processing?
SRP recognizes and binds the signal sequence as it emerges from the ribosome, pausing translation and targeting the complex to the ER membrane.
173
What are post-translational modifications (PTMs), and why are they important?
PTMs are chemical changes (e.g., phosphorylation, methylation, acetylation, glycosylation) made to proteins after translation that regulate their activity, localization, interactions, and degradation.
174
How does phosphorylation affect a protein’s function?
Phosphorylation adds a phosphate group that can change the protein’s charge, alter its conformation, and modulate its activity or binding capabilities.
175
What is ubiquitination and what is its purpose?
Ubiquitination is the covalent attachment of ubiquitin molecules to a protein, marking it for degradation by the proteasome.
176
How do deubiquitinating enzymes (DUBs) regulate protein lifespan?
DUBs remove ubiquitin tags from proteins, preventing degradation and thereby extending the protein’s functional life.
177
What is the function of the proteasome in protein processing?
The proteasome degrades ubiquitinated proteins into peptides, thereby regulating protein quality and quantity in the cell.
178
How can improper protein processing lead to disease?
Misfolded or improperly modified proteins can accumulate or lose function, potentially leading to cellular stress, dysfunction, and diseases such as neurodegenerative disorders.
179
What is recombinant DNA technology?
It involves combining DNA sequences from different sources into a single molecule (a recombinant plasmid) for use in genetic studies, protein production, or therapeutic applications.
180
What are the two key components of a recombinant plasmid?
A plasmid vector and an insert containing the gene of interest.
181
How do restriction enzymes facilitate recombinant DNA construction?
They cut DNA at specific sequences, generating fragments with either blunt or sticky ends that allow for the precise insertion of foreign DNA into a plasmid.
182
What is the advantage of using sticky ends over blunt ends in cloning?
Sticky ends have overhangs that can form hydrogen bonds with complementary sequences, increasing the likelihood of a successful ligation between the insert and the vector.
183
What role does DNA ligase play in recombinant DNA techniques?
DNA ligase catalyzes the formation of phosphodiester bonds to seal nicks, joining the insert to the plasmid vector to create a continuous DNA molecule.
184
What is bacterial transformation and why is it essential in biotechnology?
Transformation is the process of introducing a recombinant plasmid into bacteria, allowing for the amplification and production of large quantities of the plasmid.
185
What is the general approach to purifying recombinant proteins from bacteria?
Following expression, recombinant proteins are typically isolated using methods like column chromatography, which separates proteins based on specific properties such as charge, size, or affinity.
186
What are the two core components of a virus?
A virus consists of genetic material (DNA or RNA) and a protein coat called the capsid; some viruses also have a lipid envelope.
187
What are the two primary viral replication cycles?
The lytic cycle, where the virus replicates and lyses the host cell, and the lysogenic cycle, where viral DNA integrates into the host genome and replicates with it.
188
What characterizes the lytic cycle in viral infection?
During the lytic cycle, the virus commandeers the host’s machinery to synthesize viral proteins and genomes, assembles new viruses, and ultimately causes the host cell to burst (lyse).
189
How does the lysogenic cycle differ from the lytic cycle?
In the lysogenic cycle, the viral genome integrates into the host DNA, replicating passively with the host until certain conditions trigger a switch to the lytic cycle.
190
What is the role of reverse transcriptase in certain viruses?
Reverse transcriptase converts the viral RNA genome into DNA, which can then integrate into the host genome; this process is key for retroviruses.
191
What defines a retrovirus?
A retrovirus is a type of RNA virus that uses reverse transcriptase to generate a DNA copy of its RNA genome, allowing integration into the host’s DNA.
192
How can viral gene expression disrupt normal host cell functions?
Viral gene expression can hijack the host’s cellular processes, potentially interfering with cell regulation, triggering apoptosis, or even activating oncogenes.
193
What host defense mechanisms target viral infections?
Host defenses include enzymes like PKR, which detects viral double-stranded RNA, and, in prokaryotes, restriction enzymes that degrade foreign DNA.
194
In what way can viral integration lead to cancer?
Integration of viral DNA may disrupt normal gene regulation or activate oncogenes, leading to uncontrolled cell division and potentially cancer.
195
What is the primary purpose of PCR (Polymerase Chain Reaction)?
PCR is used to exponentially amplify a specific DNA sequence from a very small initial sample.
196
What are the three main steps in a PCR cycle?
Denaturation (separating DNA strands), annealing (binding of primers to target sequences), and extension (synthesis of new DNA strands).
197
What occurs during the denaturation step of PCR?
The double-stranded DNA is heated (around 96°C) to break the hydrogen bonds, resulting in single-stranded DNA.
198
What is the function of primers in PCR?
Primers are short DNA sequences that bind to the specific complementary sites on the single-stranded DNA, defining the region to be amplified.
199
What happens during the annealing step in PCR?
The temperature is lowered (typically around 55°C) so that the primers can bind (anneal) to their complementary sequences on the single-stranded DNA.
200
Describe the extension step of PCR.
At around 72°C, a thermophilic DNA polymerase adds dNTPs to the 3’ end of each primer, synthesizing a new complementary DNA strand.
201
Why is a heat-resistant DNA polymerase used in PCR?
Thermophilic DNA polymerases, like Taq polymerase, remain active at the high temperatures used during the denaturation step.
202
How does PCR achieve exponential amplification of a target DNA sequence?
With each cycle, the number of DNA molecules doubles, so after many cycles, millions to billions of copies are produced.
203
What role do dNTPs play in PCR?
dNTPs are the building blocks that the DNA polymerase incorporates to form new DNA strands during the extension phase.
204
How does PCR contribute to molecular cloning and the construction of genomic or cDNA libraries?
PCR is used to amplify specific DNA fragments, which can then be inserted into plasmids or vectors for cloning, enabling further analysis, sequencing, or protein expression.
205
What is DNA hybridization?
DNA hybridization is the process where two complementary single-stranded DNA (or RNA) molecules bind together through base pairing.
206
What is a DNA probe and how is it used?
A DNA probe is a single-stranded DNA molecule that is complementary to a target sequence; it is labeled (with radioactive or fluorescent tags) and used to detect specific sequences in a sample.
207
What is in situ hybridization?
In situ hybridization is a technique where a labeled probe is applied directly to cells or tissue sections to localize specific nucleic acid sequences within the cellular context.
208
How does DNA hybridization contribute to Southern blotting?
In Southern blotting, DNA fragments separated by electrophoresis are transferred to a membrane, where a labeled probe is used to detect fragments containing the target sequence.
209
What are the main factors that affect the stringency of hybridization?
Temperature, salt concentration, and probe length affect stringency; higher stringency conditions favor binding only between highly complementary sequences.
210
How can DNA hybridization be used in diagnostics?
It can identify genetic mutations, detect specific pathogens, or determine gene expression levels by locating specific DNA or RNA sequences.
211
Why is it necessary to denature DNA before hybridization?
Denaturation (usually by heat) separates double-stranded DNA into single strands, making the target sequence accessible for probe bindin
212
What is the difference between hybridization in Southern blotting and microarray analysis?
Southern blotting involves transferring DNA from a gel to a membrane for probe detection, while microarrays allow simultaneous hybridization of many probes to detect multiple sequences on a chip.
213
What is Sanger sequencing?
Sanger sequencing is a DNA sequencing method that uses chain-terminating dideoxynucleotides (ddNTPs) to generate DNA fragments of varying lengths for sequence determination.
214
How do dideoxynucleotides (ddNTPs) function in Sanger sequencing?
ddNTPs lack a 3’ OH group, so once incorporated into a growing DNA strand, they terminate synthesis, resulting in fragments of different lengths.
215
What is the purpose of using a mixture of dNTPs and ddNTPs in Sanger sequencing?
The mixture allows for random incorporation of chain terminators, generating a set of fragments that terminate at every possible position along the DNA sequence.
216
How are the fragments generated by Sanger sequencing separated and analyzed?
Fragments are separated by size using capillary electrophoresis or gel electrophoresis; the terminal ddNTP is then detected to read the sequence.
217
What is the role of fluorescent labeling in modern Sanger sequencing?
Fluorescently labeled ddNTPs allow each terminated fragment to be detected automatically, with different colors corresponding to each nucleotide.
218
