post midterm notes Flashcards
1
Q
who first articulated the Central Dogma and when?
A
Francis Crick in 1958
2
Q
what does DNA replication ensure?
A
that an exact copy of the species genetic info is passed from cell to cell during growth and from generation to generation
3
Q
what happens if DNA fails to replicate itself?
A
the process for meiosis and mitosis would be halted.
4
Q
true or false:
DNA replication is essential to life
A
true
5
Q
who discovered the DNA structure?
A
J. Watson, F. Crick and M. Wilkins R. Franklin
6
Q
how many base pairs are there per turn?
A
10
7
Q
how many H-Bonds are there in G C pairs?
A
3
8
Q
how many many Hbonds pair A and T?
A
2
9
Q
what kind of base pairs does DNA have?
A
complementary
10
Q
what is the most common form of DNA in living forms?
A
right-handed double helix (B-form DNA)
11
Q
how many nm between stacked bases?
A
0.34nm
12
Q
how many nm between helical turns?
A
3.4nm
13
Q
how are DNA stands made by DNA polymerase?
A
5’-3’
14
Q
what is left handed DNA?
A
Z-form DNA
15
Q
what is A-form DNA
A
- very compact form of DNA
- not found in cells
- only found in high salt concentrations
16
Q
A linear, double-stranded DNA is 10,000 bp long (10 kb).
How many complete turns of the double helix are there?
A
1,000
10,000/10
17
Q
A linear, double-stranded DNA is 10,000 bp long (10 kb).
What is the length of this molecule in μM?
A
3.4 x 1000 = 3400 nanometers or 3.4 micrometers
18
Q
A linear, double-stranded DNA is 10,000 bp long (10 kb).
How many phosphorus atoms are there, assuming one phosphorus atom per nucleotide?
A
10,000 nucleotides per strand therefore 20,000 nucleotides total
20,000 phosphorous atoms
19
Q
If there is 30% A in double-stranded DNA, how much G is there?What if the DNA was single-stranded?
A
20%
if single stranded, can’t tell composition of the rest
20
Q
how many hydrogen bonds are there in the sequence, 5’ GATC 3’3’ CTAG 5’
A
10
21
Q
Would G:C rich or A:T rich double-stranded DNA be more stable? Why?
A
G:C rich, more hydrogen bonds
22
Q
true or false:
thermotropic microbes often have very highly rich G:C DNA
A
true:
need more hydrogen bonds to stay together in harsh environments
23
Q
what were the 3 possibilities for DNA replication?
A
conservative replication
dispersive replication
semiconservative replication
24
Q
what did Meselson and Stahl do?
A
proved that the semi-conservative model was the correct copying mechanism for DNA
25
what technique did Meselson and stahl use?
cesium chloride (CsCl) equilibrium-density gradient centrifugation to separate double-stranded DNA (dsDNA) molecules of different densities
26
how does cesium chloride (CsCl) equilibrium-density gradient centrifugation world?
- permits separation of dsDNA based on density
- heavier DNA (with 15N) sediments further down the CsCl gradient
- lighter DNA (with 14N) migrates near the top of the CsCl gradient
27
which model of DNA replication applies to E. coli?
semiconservative
28
what is theta replication?
replication that occurs in most circular DNA
29
what is the product of theta replication?
2 circular DNA molecules
30
what are the 3 modes of DNA replication?
theta
rolling circle
linear chromosome
31
what is rolling circle replication?
specialized form of replication thatoccurs in the F factor and some viruses
32
is theta replication unidirectional or bidirectional?
bidirectional
33
is rolling circle replication unidirectional or bidirectional?
unidirectional
34
what are the products of rolling circle replication?
multiple circular DNA molecules
35
what is linear chromosome replication?
occurs in the linearchromosomes of eukaryotic cells
36
is linear chromosome replication unidirectional or bidirectional?.
bidirectional
37
what are the products of linear chromosome replication?
2 linear DNA molecules
38
true or false:
| linear chromosome replication has 1 origin of replication
false:
| multiple origins of replication
39
what mode of DNA replication do eukaryotic cells use?
linear chromosome replication
40
do origins of replication stay in the same spot after replication?
yes
41
what can use rolling circle replication?
small bacteria and viruses
42
what does DNA replication require?
- Magnesium (Mg2+)
- DNA dependent DNA polymerase
- 4 deoxyribonucleoside triphosphates (dNTPs)
- A template DNA to be copied
- An RNA primer
43
what does an RNA primer provide?
the 3’-OH end to initiate DNA synthesis by DNA polymerase
44
true or false:
| DNA is alwayssynthesized in the 5’ to 3’ direction
true
45
true or false:
| Newly-synthesized DNA strand is complementary and anti-parallel tothe parent strand
true
46
Howard DNA strands held together?
by hydrogen bonds between complementary bases
47
A fragment of partially double-stranded DNA has the structure (5’ denotes the 5’ monophosphate; 3’ denotes the 3’ OH group),
5’ AGCTAGTTATTACG 3’
TCAATAAT
If this DNA was used as a template for replication, whichnucleotide would be incorporated first?
A
goes from 5' to 3'
will extend to the left, right side will not extend anymore
48
where does chain cleavage lave the alpha phosphate group?
attached to 5' or 3' carbon
49
where is DNA synthesis continuous?
on the leading strand
50
where is DNA discontinued?
on the lagging strand
51
what are Okazaki fragments?
The short DNA fragments produced by discontinuousDNA synthesis
52
how many dnaA binding sites does E. coli have?
4
53
where does helices bind?
the strand that will express lagging strand synthesis
54
how does DNA helices unwind DNA?
in the 5’ to 3’ direction: travels on the lagging strand ahead of the replication machinery
55
what is unwound ss DNA coated with?
single-strand binding protein to keep DNA single-stranded
56
what does helicase-induced unwinding of the double helical DNA cause?
DNA ahead of the helicase to be overwound producing positive supercoils that would stop replication.
57
what nicks the DNA and releases the positive supercoils in bacteria?
specialized DNA topoisomerase II (DNA gyrase)
58
what does DNA primase synthesis?
short RNA primer that provides the 3’ OH end for DNA polymerase to begin DNA synthesis
59
how many DNA polymerase are there in E. coli?
5
60
what are the main DNA polymerases in E. coli?
I, III
| Chromosomal DNA replication-replicative polymerases
61
what are DNA polymerase II, IV, V in E. coli?
DNA repair function
62
what are the activities of DNA polymerase I?
- aids in removal of RNA primers
- has 5’ to 3’ polymerase and 5’ to 3’ exonuclease activity
- Proofreading: has 3’ to 5’ exonuclease activity
- not highly processive; short tract synthesis
63
what are the Main activities on DNA polymerase III?
- main replicative polymerase; highly processive
- has 5’ to 3’ polymerase activity
- lacks 5’ to 3’ exonuclease activity
- Proofreading: has 3’ to 5’ exonuclease activity
64
what is the beta sliding clamp?
a ring-shaped polypeptide that encircles the DNA and interacts with DNA polymerase III to enhance processive DNA synthesis
65
What does ss DNA binding protein do?
keeps the unwound strand in an extended form for replication
66
what direction does all DNA synthesis occur in?
5' to 3'
67
how fast is DNA synthesis in E. coli?
4000 base pairs added per second
68
what does the replication mechanism require?
- topoisomerase
- helicase
- Single-strand DNA binding protein (SSB)
- DNA primase
- DNA polymerase III (plus βclamp)
- DNA polymerase I
- DNA ligase
69
what is the frequency for replicative polymerase?
30,000 nt/min
70
frequency of errors in replicative polymerase
1 error/ 10^10 nt added
71
describe eukaryote RNA primers and Okazaki fragments
shorter
72
when does DNA replication occur in eukaryotes?
S phase
73
what does Pol epsilon do in eukaryotes?
performs leading strand replication
74
what does Pol delta do in eukaryotes?
performs lagging strand replication
75
what does Pol alpha do in eukaryotes?
synthesizes has primase activity
76
true or false:
| eukaryotes have Bidirectional replication from multiple origins of replication on each chromosome
true
77
what happens to the nucleosomes in eukaryotes?
need to be removed from parental DNA andproperly re-assembled on newly-synthesized DNA
78
describe telomeres in eukaryotes
shorten at each round of eukaryotic replication
79
true or false:
| disassembly and Assembly of Nucleosomes is Tightly coupled and rapid during DNA synthesis
true
80
what is the telomere problem?
chromosome end will be degraded causing chromosome shortening during every round of replication
81
what does telomerase activity do in eukaryotes?
extends eukaryotic chromosome ends in replication
82
true or false:
| telomerase does not resolve the terminal primer problem
false
83
true or false:
| Most human somatic cells have high telomerase activity
false:
| low
84
what are shorter telomeres associated with?
cellular senescence and death
85
what are diseases causing premature aging associated with?
short telomeres
86
true or false:
| cancer cells have been found to have high telomerase activity
true:
| thought to promote their growth
87
what are the 2 olecular biology techniques that are based upon fundamental knowledge of DNA replication?
1. DNA sequencing
| 2. Polymerase chain reaction
88
what does DNA dependent RNA polymerase do?
moves along DNA to produce RNA
| transcription
89
Describe what RNA contains
- has a ribose sugar (bears an –OH group on its 2’ carbon)
| - contains the base uracilin place of thymine
90
what kind of structure does RNA have?
tertiary structure
| potential to be quaternary structure
91
what does structure in DNA affect?
how molecules bind to it
92
true or false:
| transcription and translation happen at the same time in prokaryotes
true
93
true or false:
| prokaryotes, the coding region of a gene is often a single, continuous unit
true
94
true or false:
| tanscription, translation and mRNA degradation often occur at different times in prokaryote
false:
| usually occurs simultaneously
95
what are exons?
protein coding segments
96
what are introns?
intervening (non-coding) segments
97
what happens when anscription and translation are not coupled-transcripts ?
made and processed in the nucleus and must be transported to the cytoplasm for translation
98
what is messenger RNA (mRNA)
intermediates that carry genetic information from DNA to the ribosomes
99
what is Transfer RNAs (tRNAs)
adaptors between amino acids and the codons in mRNA.
100
what is ribosomal RNAs (rRNAs)
structural and catalytic components of ribosomes
101
what is small nuclear RNAs (snRNAs and snoRNAs)
spliceosomes and rRNA, tRNA modification, respectively.
102
what are micro RNAs (miRNAs, siRNA and Crispr RNA)
short RNAs that block expression of complementary mRNAs
103
what is long noncoding RNAs
long RNAs that regulate gene transcription
104
who has a nucleus; eukaryotes or prokaryotes?
eukaryotes
105
which RNAs do not get made into proteins?
tRNAs, RNAs, nRNAs and snoRNAs, iRNAs, siRNA and Crispr RNA, long noncoding RNAs
106
what is the most frequent RNA in a cell?
ribosome (>80%)
107
what is the only kind of RNA that encodes proteins?
Messenger RNA (mRNA)
108
how is RNA synthesized?
in the 5’ to 3’ direction using the 3’ to 5’DNA template strand
109
true or false:
| RNA and DNA strands are parallel
false:
| antiparallel
110
how can transcription utilize the 3’ to 5’ template
either single strand of the double-stranded DNA (dsDNA), but always occurs in the 5’ to 3’ direction
111
true of false:
| genes can overlap in 5’ to 3’ transcription
true
112
what is the DNA template?
4 ribonucleoside triphosphates (rNTPs)A, U, C, G
| RNAn+ rNTP ---> RNAn +1+ PPi
113
what does transcription use instead of deoxynucleoside triphosphate that are used in DNA replication?
ribonucleoside triphosphate
114
```
A segment of mRNA has the sequence, 5’AUCCUGA 3’Which single-stranded DNA was it transcribed from?
A.5’ TAGGACT 3’
B.5’ UAGGACU 3’
C.3’ TAGGACT 5’
D.3’ UAGGACU 5’
E.More than one of the above
```
C.3’ TAGGACT 5’
115
what are some general features of RNA synthesis?
- Similar to DNA Synthesis except
–The precursors are ribonucleoside triphosphates (rNTPs)
–Only one strand of DNA is used as a template.
–RNA chains can be initiated de novo (no primer required)
116
how will RNA related to DNA?
complementary to the DNA template strand and identical to the DNA non-template strand
117
what is RNA synthesis catalyzed by?
RNA polymerases and proceeds in the 5’ to 3’ direction
118
what are the 3 stages of transcription?
1. initiation
2. elongation
3. termination
119
describe initiation of transcription
- RNA polymerase binds, unwinds and joins first 2 nucleotides.
- initiation of RNA synthesis DOES NOT require a prime
120
describe elongation of transcription
- complementary nucleotides continue to be added during the elongation process.
- localized DNA unwinding ahead of RNA polymerase generates a “transcription bubble”.
- transcription bubble moves with theRNA polymerase and the unwound DNA rewinds behind it
121
describe termination of transcription
- stops when RNA polymerase reaches the “terminator”region of the gene
- the newly-synthesized RNA together withRNA polymerase are released.
122
what are the 2 important sequence elements in a typical E. coli promoter?
- the -35 element to which σfactor binds
| - the -10 element, which due to it very A/T rich content is prone to unwinding
123
what do promoter recognition require in the initiation of transcription?
the RNA polymerase holoenzyme
124
what does the sigma factor recognize and bind to in the initiation of transcription?
the -35 element, thus positioningthe RNA polymerase at the promoterto begin transcription
125
what is the pribnow nob?
- 35 sequence: 5’ TTGACA 3’
| - 10 sequence: 5’ TATAAT 3’
126
where does transcription initiate?
about 5-9 base pairsdown from the end of the -10 sequence
127
what is the 5' end of the RNA usually?
purine
128
when does elongation occur?
when Sigma factor is released and RNA polymerase begins to move along the 3’ to 5’ DNA template strand
129
what occurs as RNA polymerase moves along the DNA template?
A localized region of unwinding called the “transcription bubble”
130
what is removed by topoisomerases?
Positive supercoils formed in the double-stranded DNA ahead of the advancing RNA polymerase
131
true or false:
| RNA polymerase has both helix unwinding and rewinding activities
true
132
what does Weak H-bonding at U:A residues allow?
mRNA release from DNA whenRNA polymerase pauses at terminator
133
what are Puffs in Drosophila polytene salivary chromosomes?
sites of localized unwinding due to gene transcription
134
true or false:
| there are specific promoters for genes transcribed by Pol. I, Pol. II OR Pol. III
true
135
what do accessory proteins do in eukaryotes?
recognize each specific type of promoters (through interaction with DNA sequences) and recruit the appropriate polymerase to begin transcription
136
which has more complex promoters: eukaryotes or prokaryotes?
eukaryotes
137
what does eukaryotic initiation in the case of Pol II promoters involve?
step-wise assembly of general Transcription Factors of Pol. II (TFII A, B, D, E, F and H)
138
what does the preinitation complex do?
is sufficient to initiate transcription.
139
what does more complex transcriptional regulation involve?
a multi-subunit complex called “Mediator” that permits interactions with other activator proteins bound to upstream/downstream regulatory regions or enhancer sequences
140
what happens during elongation in eukaryotes?
- many of the general transcription factors remain at the promoter providing for quick re-initiation with a new Pol. II.
- an ~8 nucleotide “transcription bubble”is generated by RNA:DNA binding.
- this together with DNA unwinding, ensures that the free RNA 3’-OH terminus is available for new rNTP addition.
141
what is alpha amanitin?
a molecules made from the death cap mushroom
142
what is alpha amanitin known as?
a known potent inhibitor RNA polymerase
143
what does termination in eukaryotes involve?
- cleavage of the pre-mRNA and 5’ to 3’ degradation of the remaining RNA by the Rat1 exonuclease.
- transcription terminates when Rat1 reaches RNA polymerase
144
describe the coding region in prokaryotes
coding region of a gene is not interrupted: the sequence of the gene is co-linear with the amino acid sequence of the protein
145
true or false:
| the prokaryotic messenger RNA sequence corresponds to the gene from which it was transcribed
true
146
what is the Shine-Dalgarno sequence and what is it involved in?
5’ UAAGGAGGU 3’ is involved in the initiation of translation
147
when was the presence of intron in eukaryotic genes discovered?
1970s
148
what is required to form the mRNA that will be translated into a polypeptide?
the removal of introns along with additional RNA processing steps
149
what are the 3 main processing steps in eukaryotic nuclear pre-mRNA?
A. addition of 7-methyl guanosine cap
B. addition of polyA tail
C. removal of introns
150
describe A. addition of 7-methyl guanosine cap
- Linked to pre-mRNA by a unique 5’-5’ phosphate linkage
| - addition of 7-MG cap occurs early in the elongation process
151
describe . Addition of PolyA tail
- pre-mRNA is cleaved and then, a long string of “A” residues is added by Poly A polymerase
- eukaryotic pre-mRNA is cleaved 11-30 nt following the 5’ AAUAAA 3’ sequence in the pre-mRNA.
- Poly A polymerase adds a string of ~200 A residues at the cleaved end
152
describe C. removal of introns
- introns in pre-mRNA are removed by a specialized process called“RNA splicing”
- removal of introns must be precise in order to properly fuse the 3’ end of one exon to the 5’ end of the next exon.
153
what does every intro have?
two conserved sequences that are required for its removal
154
what are the 2 conserved sequences that are required for an introns removals?
A. 5’ and 3’ splice sites: “GU and AG” sequences, respectively.
B. Intron Branch point: conserved “A” residue
155
what is a spliceosome?
- RNA/protein structure
- Five small nuclear RNAs (snRNAs): designated U1, U2, U4, U5, and U6
- The snRNAs associate with about 40 small proteins to form small nuclear ribonucleoproteins (snRNPs)
156
what are the snRNPs that are assembled to form a complete spliceosome?
U1, U2, U4/6 and U5
157
what does lariat formation involve?
a unique linkage between the 5’phosphate of the “G” and the2’ OH of the “A"
158
how are introns of nuclear pre-mRNA transcript carried out?
by splicosomes
159
how are introns of some rRNA precursors removed?
autocatalytically by reaction of the RNA molecule itself
160
how are introns of tRNA precursors excised ?
by precise endonucleolytic cleavage and ligation reactions.
161
how can 1 gene make many proteins?
presence of introns allows for alternative-modes of splicing and 3’ end processing that can generate protein diversity
162
how many genes is it estimated that alternative processing occurs in for humans?
>90%
163
true or false:
| genetic info is usually altered in the mRNA intermediary
false
164
what are the 3 mechanisms that RNA editing can occur?
- Changing the structures of individual bases (i.e., tRNAs, rRNAs).
- Modification of mRNA by endogenous guide RNAs
- Inserting or deleting uridine monophosphate residues
165
what is an apoplipopotein?
blood proteins that carry lipids (fatand cholesterol) in the bloodstream
166
what enzyme helps with RNA editing of ApoB?
Cytidine deaminase
167
what does RNA editing f ApoB by the enzyme Cytidine deaminase do?
changes “C” to “U” converting a normal glutamine codon (CAA) to a termination codon (UAA), which truncates the protein and gives it a different function with respect to lipid binding
168
what does the anticodon of the tRNA base pair bind with?
the codon of mRNA
169
where is an amino acid covalently attached to?
the 3' end of tRNA
170
what do tRNAs often contain?
modified nucleotides through the action of tRNA modifying enzymes and they undergo processing events that remove small introns
171
what are ribosomes composed of?
a large and a small subunit that are assembled from many different proteins and rRNAs
172
what is a ribosome?
an “RNA machine” with key roles in protein synthesis, including the formation of peptide bonds between amino acids
173
what is the nucleolus?
site of eukaryotic rRNA synthesis and ribosome assembly
174
where does rRNA synthesis and ribosome assembly occur in prokaryotes?
in the cytoplasm
175
what do enzymes do to rRNA transcripts?
modify and trim precursor rRNA transcripts to mature forms in the cell
176
describe snRNAs
- act complexes with proteins.
| - play roles in post-transcriptional processing of RNA, such as splicing
177
describe snoRNAs
- act in complexes with proteins.
| - guide the enzymatic chemical modifications of ribosomal RNAs, transfer RNAs and small nuclear RNAs
178
describe small micro RNAs in eukaryotes
- act as short (~22 nt), single-stranded RNAs that bind to complementary sequences in mRNA.
- produced by cleavage of mRNAs, RNA transposons, and RNA viruses.
- regulate and control gene expression in different ways
179
describe small micro RNAs in prokaryotes
- Crispr RNA- encoded by DNA sequences found in prokaryotic genomes.
- Works in association with the prokaryotic Cas9 nuclease to cleave foreign DNA that might happen to enter a host cell (prevents incorporation of foreign DNA into the host genome
180
where are long noncoding RNAs knows to function?
eukaryotic cells
181
what is the signifiants of long noncoding RNAs?
about 80% of the mammalian genome consist of non-protein coding RNAs
182
what is the function of long noncoding RNAs?
- regulate and control gene expression at the level of transcription or translation by binding mRNA or sequestering micro-RNAs that control gene expression.
OR
- bind and recruit proteins involved in DNA modification
183
what is sickle cell anemia caused by?
a single aminoacid change (Glu to Val) in one of the protein chains that make up hemoglobin in red blood cells
184
what is an mRNA transcript?
the RNA copy of the template DNA strand of the gene
185
what is the purpose of translation?
to decode the mRNA and make the functional protein product of the gene
186
true of false:
| Transcription, translation and mRNA degradation often occur simultaneously in prokaryotes
true
187
where does transcription occur in eukaryotes?
the nucleus
188
where does mRNA translation occur in eukaryotes?
the cytoplasm
189
what do The sequence of base pair triplets in the coding region of a gene specify?
a colinear sequence of amino acids in its polypeptide product
190
what did Beadle and Tatum: in the 1930s do?
through genetic analysis of nutritional mutants in the fungus Neurospora, they discovered that one gene encoded one discrete polypeptide
191
what did Charles Yanofsky and colleagues: in the 1960s do?
through mutational/biochemical analysis, they discovered that the sequence of nucleotide triplets in the trpA gene of E. coli corresponded to the sequence of amino acids in the TrpA protein
192
what are proteins made of?
polypeptides
193
what is a polypeptide?
a long chain of amino acids
194
what are amino acids made of?
a free amino group, a free carboxyl group, and a side group (R
195
what are amino acids joined by?
peptide bonds
196
what are peptide bonds?
The carboxyl group of one amino acid is covalently attached to the amino group of the next amino acid.
197
describe primary structure of proteins
Linear arrangement of amino acids
198
describe secondary structure of proteins
Determined by the spatial organization of amino acids
199
describe tertiary structure of proteins
determined by the overall folding of the complete polypeptide
200
describe quaternary structure of proteins
in some proteins, more than one polypeptide interacts to make a functional protein
201
what is a singlet code?
since only 4 bases, only 4 codons specified...not enough for 20 amino acids
202
what is a doublet code?
would specify only 42 = 16 possible codons... not enough for 20 amino acids
203
what is a triplet code?
would specify, 43 = 64 possible codons... sufficient for synthesis of the 20 amino acids IF some amino acids were specified by more than one codon
204
what kind of code is the genetic code?
triplet code
205
what did arshall Nirenberg, Gobind Khorana, Philip Leder and colleagues play essential roles in?
deciphering the genetic code
206
how was the genetic code deciphered?
1. synthetic mRNAs of repeated sequence (homopolymers) were tested in vitro for protein synthesis: produced homopolypeptides
2. Mixed mRNAs (random copolymers) were tested in same way: produced polypeptides with different amino acids.
207
what did Nirenberg and Leder demonstrate in further experiments?
short mRNAs of known sequence stimulated the binding of ribosomes and the corresponding amino-acid bound tRNA
208
true or false:
| all Aino acids have 1 codon?
false:
| some have more than one
209
how many stop codons are there?
3
210
how many start codons are there?
1
211
why is there more than one codon for some amino acids?
Oftentimes, the base in the 3rd codon position can be changed and still specify the same amino acid: degeneracy
212
true or false:
| Base-pairing between mRNA codons and aminoacyl tRNAs is “anti-parallel"
true
213
describe the wobble hypothesis
- stringent base pairing between the codon in mRNA and the anti-codon intRNA only occurs for the first two bases of the codon
- base-pairing at the third base of the codon is less stringent allowing “wobble”or flexibility at this position
214
what does the wobble hypothesis explain?
how a single tRNA can respond to two or more codons
215
what is at the wobble position?
ydrogen bondingand the non-standardbase-pairing
216
what is inosine (I)?
a modified guanine/adenine derivative
217
what are ribosomes made up of?
polypeptides (> 50) and ribosomal RNA (rRNA) molecules (3-4)
218
what are macromolecules of translation made up of?
- ribosomes
- Amino-acid Activating Enzymes (20)
- tRNA Molecules (40-60)
- Soluble proteins
219
what is a ribosome?
an “RNA machine” with key roles in protein synthesis, including the formation of peptide bonds between amino acids
220
what are tRNAs?
adapters between amino acids and the codons in mRNA
221
who and when was tRNA populated by?
Francis Crick in 1958
222
what does he anticodon of the tRNA base pair with?
codon of mRNA
223
where is the AA attached to tRNA?
covalently attached to the 3' end
224
how much tRNA synthetase does each ell contain?
1 for each amino acid
225
what does tRNA synthetase catalyze?
formation of aminoacyl tRNA
226
describe inititation f translation?
- mRNA, large and small ribosomal subunits, initiation factors (IF1-3) and GTP are all required to form the initiation complex
- 16S rRNA is a component of the 30S ribosomal (small) subunit
- pairing between these sequences is involved in the formation of the mRNA/30S ribosomal subunit initiation complex
227
what are the sites in the ribosome?
A - Aminoacyl site
P- Peptidyl site
E- Exit site
228
what is the difference between the amino group of the methionine on the initiator tRNA in eukaryotes and prokaryotes?
is not formylated methionine in eukaryotes: simply MET
229
what is there none of at the translation start site in eukaryotes?
no Shine-Dalgarno/AUG translation start site
230
what do eukaryotes do to find the AUG initiation codon?
the initiation complex scans the 5’ end of the mRNA
231
what does the Kozak sequence do in eukaryotes?
influences the efficiency of which AUG in the vicinity of the 5’ end is used to start translation
232
what is the Kozak sequence?
at the 5’ end of the mRNA, the Kozak sequence is,5’- GCC (A or G) CC AUGG 3'
233
what does the poly(A) tail of the eukaryotic mRNA interact with?
the mRNA 5’ cap structure via protein a cap-binding complex (CBC) to promote to promote translation initiation
234
describe elongation of translation
- An aminoacyl-tRNA binds to the A site of the ribosome
- The amino acid is transferred from the tRNA in the P site to the tRNA in the A site by the formation of a peptide bond
- The ribosome translocates along the mRNA to position the next codon in the A site.
- This results in the polypeptide-tRNA being translocated from the A site to the P site
- The uncharged tRNA is translocated from the P site to the E site and removed.
- A new, aminoacyl-tRNA agains binds in the A site and the cycle is repeated
235
when does termination of translation occur?
when a stop codon enters the A site of the ribosome.
236
what are the stop codons?
UAA, UAG, and UGA.
237
what happens when a stop codon is reached?
- a release factor (RF) binds to the A site.
238
what does release factor 1 recognize?
UAG and UAA
239
what does relate factor 2 recognize?
UAA and UGA
240
what did e discovery of ribosome structure permit?
detailed molecular analysis of how ribosomes work during protein synthesis
241
what does chloromycetin inhibit?
formation of peptide bonds
242
what does erythromycin inhibit?
translation of mRNA along ribosome
243
what does neomycin inhibit?
interactions between tRNA and mRNA
244
what does streptomycin inhibit?
initiation of translation
245
what does tetracycline inhibit?
binding of tRNA to ribosome
246
what does papomomycin inhibit?
validation of mRNA-tRNA match
247
what do mRNA specialized mechanisms do?
- eliminate mRNAs with nonsense mutations.
- eliminate mRNAs in which a stalled ribosome cannot complete proper translation
- eliminate mRNAs that are damaged by chemicals, unusual secondary structures, etc
248
what do Molecular chaperones function to assist ?
proper foldingof newly-synthesized proteins and are often associatedwith the ribosome during translation.
249
what are the steps of translation?
1. polypeptide chain initiation
2. chain elongation
3. chain termination
250
what is the Shine-Dalgarno sequence?
5' AGGAGG 3'
251
who helped with our understanding of how the ribosome works?
Venkatraman Ramakrishnan,
Thomas A. Steitz,
Ada E. Yonath
252
what is Xeroderma pigmentosum?
lack of nucleotide excision repair of UV-induced lesions
(DNA repair pathway defect)
- develop skin cancer if pyrimidine dimers don't get removed
253
true or false:
| changes in DNA structure and errors in replication dooccur
true:
| just rare
254
how is the vast majority of cellular DNA damage corrected?
by efficient DNA repair mechanisms.
255
what happens what DNA damage is not corrected?
an inherited changein genetic informationmay occur and this is a “mutation”
256
what is a mutation?
- A heritable change in the sequence of an organism’s genetic material-the mutation may alter the phenotype of the organism.
- The process by which genetic change occurs
257
what is a mutant?
An organism that carries one or more mutations in its genetic material
258
what do mutations occur in?
in all organisms with genetic material, from viruses to humans
259
true or false:
| Mutation is the source of all genetic variation.
true
260
what happens during meiosis regarding mutations?
recombination between homologous chromosomes rearranges genetic variability into new gene combinations
261
what is a somatic mutation?
occur in somatic cells; it will occur only in the descendants of that cell and will not be transmitted to the progeny.
262
what is a germinal mutation?
occur in germ-line cells and will be transmitted through the gametes to the progeny.
263
what were the original mutations that occurred in fruit trees?
- somatic mosaics
| - vegetative propagation allowed the somatic mutation to be perpetuate
264
true or false:
| somatic mutations can lead to cancer
true
265
what is the expanding of nucleotide repeats known as?
“dynamic mutation” because the nucleotide repeat copynumber can expand or contract dramatically in each cell
266
what occur at localized sites in DNA-often referred to as "point mutations"?
1. Base substitution
2. Frameshift mutation
3. Tautomeric shifts
267
what is gross chromosomal rearrangement?
A change in chromosomenumberor structure
268
describe base substitution
replacing one base with another
269
describe transition base substitution
replaces a pyrimidine with another pyrimidine or a purine for another purine
270
describe transvehrsion base substitution
replaces a pyrimidine with a purine or a purine with a pyrimidine
271
how many different base substitutions can occur in DNA?
12
272
describe frameshift mutations
insertions or deletions of 1-2 base pairs alter the reading frame of the gene detail to the site of mutation
273
true or false:
| protein sequence is fairly similar after the frameshift mutation
false:
| changes dramatically
274
what is a tautomeric shift?
movement of H atoms from one position in a purine or pyrimidine base to another
275
are tautomeric shifts common or rare?
rare:
can occur spontaneously during DNA replication where they alter DNA base pairing. Cause some spontaneous mutations
(A:C and G:T base pairs)
276
when can bases form A-C and G-T base pairs?
When the bases are in their rare enol or imino states
277
what is a G:C to A:T base pair change:?
transition mutation
278
what are the normal forms of the DNA bases?
amino forms and keto forms
279
what does the mechanism of expansion involve?
DNA replication
280
what is a forward mutation?
a genetic alteration that changes the wild-type phenotype to mutant
281
what is a reverse mutation?
changes the mutated site back to normal, hence reversing the mutant back to the wild-type phenotype
282
what is a missense mutation?
a base substitution that results in an aminoacid change in the protein
283
what is a nonsense mutation?
a base substitution that changes a sensecodon to one of the three nonsense (stop) codons (UAG, UGAor UAA)
284
what is a silent mutation?
a base substitution at the 3rd codon position that changes the codon to one still specifying the same amino acid
285
what is a neutral mutation?
a missense mutation in which the amino acid is changed to one of a similar chemical type
286
what is a loss-of-function mutation?
he result of mutations that cause complete or partial loss of normal protein function
287
what is a gain-of-function mutation?
the result of a mutation that causes the cell to produce a protein or gene product whose function is not normally present
288
what is a conditional mutation?
expressed only under certain conditions (i.e., a temperature-sensitive mutation-only observed at elevated temps)
289
what is a lethal mutation?
causes premature cell death
290
what is a suppressor mutation?
a second site mutation that hides or suppresses the effect of the first mutation.
291
true or false:
| suppressor mutations can be within the same gene or different gene
true:
same - intragenic suppressor
different - intergenic suppressor
292
true or false:
| spontaneous mutations are frequent
false:
If we assume a typical gene is ~103nucleotide pairs, then the mutation rate varies from ~10–4to 10–7 per gene per generation
293
how much can treatment of bacteria with mutagens increase mutation frequency?
>1% per gene
294
what are mutations that occur under normal circumstances a result of?
internal factors are called “spontaneous” mutations
295
what are mutations that occur as a result of external factors called?
“induced” mutations
296
what is most DNA damage caused by?
internal factors generated by normal metabolic processes
297
true or false:
| we have several dedicated DNA repair systems
true
298
what does spontaneous DNA damage result from?
- DNA replication errors
- DNA replication pausing
- Endogenous chemical reactions
299
what are examples of DNA replication errors?
- tautomeric shifts
- wobble-induced base misfiring
- strand slippage during replication
300
describe how wobble-induced base misfiring occurs
flexibility in base-pairing (or wobble) can result in non-standard G-T and C-A base pairs, which lead to transition mutations
301
what is strand slippage during replication?
occurs in repeated DNA sequences. Also, misalignments during recombination
302
what is DNA replication pausing?
- replication stalling at a DNA nick
- unusual DNA structure or bulky lesion can generate broken DNA such as DNA ends or double-strand breaks (DSB), which can be lethal or mutagenic
303
what are the 4 types of endogenous chemical reactions?
1. depuration
2. deamination
3. oxidation
4. alkylation
304
what is depuration?
spontaneous loss of a purine base from a nucleotide through hydrolysis of the glycosidic bond
- generates transition or transversion mutations
305
which is more frequent: loss of a purine or pyrimidine base?
loss of a purine
| 10,000 cell/day vs. 500 cell/day
306
what is deamination?
- spontaneous loss of -NH2 group of DNA base
| - causes transition mutation
307
what is oxidation?
- endogenous reactive oxygen species (ROS) damage DNA
| - Can produce oxidized bases, such as 8-oxoG, which frequently mispairs with C or Ato producetransversion mutations
308
what is alkylation?
endogenous alkylating agents can add methyl groups to DNA bases
- produce transition mutations
309
what mutagens induce DNA damage?
- chemical agents
| - radiation
310
when were induced mutagens first discovered and by who?
1927 by Hermann Muller
311
what did Muller's experiment entail?
can induce mutations in the fruit fly Drosophila using X-rays: through studies of X-linked mutations.
312
what are the categories of mutations caused by chemical agents?
- mutagenic to both replicating and non replicating DNA
| - mutagenic only to replicating DNA
313
what are alkylating agents?
- mutagens that great with DNA bases and add methyl or ethyl groups
- directly or indirectly can induce transitions, transversions, frameshifts and chromosomal aberration
314
what is nitrous acid?
- deaminating agent.
- Removes amino (–NH2) groups from DNA bases A, C and G.
- Cause transition mutations
315
what is hydroxyl amine?
- hydroxylating agent
- -hydroxylates the amino (-NH2) group of cytosine causingthe modified base to pair with adenine after replication
- transition mutation
316
what are the 2 common base analogs?
- 5-bromouracil (5-BU) (resembles T)
| - 2-aminopurine (2-AP) resembles A or G)
317
what do base analogs do?
Incorporated into DNA during replication and when rare tautomers arise will eventually cause transition mutation
318
describe acridines
ercalation of an Acridine dye causes frameshiftmutations during DNA replication
319
what are the 2 main types of radiation that cause mutations?
- Ultraviolet (UV) light induces mutations through excitation.
- X-rays (and shorter wavelengths) induce mutations through ionization
320
at peak sunlight, what does each human skin cell acquire?
4500 thymine dimers and UV-induced photoproducts per hour!
321
what does ionizing radiation cause?
nicks and DSB in chromosomes
322
what can faulty repair of DNA breaks by recombination cause?
gross chromosomal rearrangements such as deletions, duplications, inversions, and translocations
323
NA is constantly being damaged by endogenous (mostly) and exogenous agents.-in spite of this, the rate of mutation in cell is remarkably low
why?
we have a number of very potent mechanisms for repairing DNA damage
324
what are our DNA repair mechanisms?
- Direct reversal of DNA damage
- Excision repair (base excision and nucleotide excision)
- Mismatch repair
- Recombination
- Translesion DNA polymerases (Error-prone repair)
325
how does direct reversal of DNA damage work?
1. Light-Dependent Repair: Direct repair of thymine dimers by the enzyme, photolyase (called, photoreactivation: only found in prokaryotes)
2. Enzymatic removal of alkyl groups from DNA bases
3. Ligation of single-stranded nicks in DNA.
326
what are the 2 kinds of excision repair?
- base excision repair
| - nucleotide excision repair
327
what are the common features of excision repair?
- A DNA repair endonuclease or endonuclease-containing complex recognizes, binds to, and excises the damaged base or bases.
- A DNA Polymerase fills in the gap, using the undamaged complementary strand of DNA as a template.
- DNA ligase seals the break (nick) left by DNA polymerase
328
how does base excision repair (BER) work?
- a dedicated repair mechanism that specifically recognizes and repairsDNA bases damaged by deamination, alkylation or oxidation.
- found in both prokaryotes and Eukaryotes
329
how does nucleotide Excision Repair (NER) work?
NER acts to remove thymine dimersand other bulky forms of DNA damage
330
what do mutations of human NER genes cause?
Xeroderma pigmentosum
331
how does mismatch repair (MMP) work?
- recognizes a mismatched base in the newly-replicated DNA strand through identification of the hemi-methylated GATC sequence
- compares old and new strands based on methylation status of “A” in the sequence,
- After detection, an exonuclease removes a portion of the newly-synthesized strand that includes the incorrect base.
- DNA polymerase fills in the gap and ligase seals the nick
332
wha do defects in the human mismatch repair result in?
cancer
333
how much of colorectal and endometrial cancers are hereditary and cause by mutations in human mismatch repair genes?
10%
334
what are the 2 types of recombination?
- homologous recombination (HR)
| - non-homologous end joining (NHEJ)
335
how does homologous recombination work?
- often occurs during or after DNA replication.
- If one sister chromatid suffers a DSB, it can be repaired using the identical (unbroken) sister chromatid.
- usually occurs in the S/G2 phases of the eukaryotic cell cycle
- used to ensure proper chromosome separation (dysjunction )during meiosis
336
how does non-homologous end joining work?
- uses an entirely different set of proteins to repair DSB.
| - available throughout the rest of the eukaryotic cell cycle
337
what is Post-replication Recombination Repair?
- involves pairing and strand exchanges between DNA duplexes
- provides a mechanism for T:T to be by-passed so that replication can continue.
- repairs double-stranded breaks in chromosomes
338
what is the SOS response?
In the event that DNA is heavily damaged by mutagenic agents, a DNA damage response is activated
339
true or false:
| in bacterial cells, an increase in mutation = adaptation to environment
true
340
what do pyrimidine dimers and other bulky DNA lesions block resulting in cell death if replication cannot continue?
replicative polymerases
341
what are transposable elements?
segments of DNA capable of moving from one location in a chromosome to another, or even to a different chromosome
342
what are the 3 categories of transposable elements?
1. cut and paste transposons
2. replicative transposons
3. retrotranspons
343
what are cut and paste transposons?
element is physically cut out of one site in a chromosome or plasmid and pasted into a new site
- DNA transposon
- excision and insertion catalyzed by a transposase
- found in both prokaryotes and `eukaryotes
344
what are replicative transposons?
element is replicated with one copy inserted ata new site and one remains at original site
- DNA transposon
- requires transposase
- only found in prokaryotes
345
what are retrotansposons?
- DNA copy of element made by reversetranscription from its RNA and then inserted into a new chromosomal site
- only found in eukaryotes
- two kinds: Retrovirus-like elements and retroposons
346
true or false:
| Bacterial transposons move within and between chromosomes and plasmid
true
347
what is a gene that encodes transposase?
a protein required for transposition to occur
348
what are terminal inverted repeats?
identical, or nearly identical inverted sequences at both ends of element
349
what are target site duplication?
short directly repeated sequences at both ends -result from staggered cleavage of the double-stranded DNA at the site of insertion
350
what are the simplest bacterial transposons?
Insertion Sequences (IS elements)
351
what are IS elements?
- compactly organized and contain only genes whose products are involved in transposition.
- inverted terminal repeats are found at the ends.
- Some encode transposase
352
what does insertion of an IS element cause?
traget site duplication
353
what may happen when a particular IS element if found on both a plasmid and a chromosome?
homologous recombination may occur inserting the plasmid into the chromosome
354
what is the R-determinant?
antibiotic resistance gene
355
what is the RTF component?
resistance transfer factor
356
when are composite transposons created?
when two IS elements insert near each other, “capturing” a DNA sequence
357
what can IS element excision mobilize?
the captured DNA, which may mobilize antibiotic resistance genes
358
where are replicative transposons found?
bacteria
359
true or false:
| Tn3 elements are larger than the simple IS element
true
360
what do Tn3 elements often contain?
additional genes that are not involved in transpositio
361
how were transposonsable elements discovered?
by analyzing genetic instabilities in maize
362
what did Barbara McClintock do?
discovered transposons
363
how much of our genome is made up by transposable elements?
44%
