Week 2 Flashcards
1
Q
What underlies the production of all cellular proteins?
A
Translation of mRNA into polypeptides.
2
Q
How are 20 amino acids encoded in the genetic code?
A
Four bases in RNA/RNA encode 20 amino acids.
DNA RNA Protein
4 nt 4 nt 20 AA
3
Q
What are the building blocks of proteins?
A
Amino acids
4
Q
What are non-polar amino acids?
A
are hydrophobic (often in the core of proteins, where they avoid water)
5
Q
What are the characteristics of amino acids with polar side chains?
A
are hydrophilic (tend to be on outside of proteins) By interacting with water they make proteins soluble
6
Q
What does the amino acid sequence lead to?
A
leads to proteins having a 3D structure
7
Q
What do proteins make up?
A
Hair and nails Blood Brain and nerves Enzymes cellular construction workers. Antibodies Cellular messengers Muscles
8
Q
What does a triplet code offer?
A
64 combinations
9
Q
What are the properties of the genetic code?
A
Triplet
Non-overlapping
Non-ambiguous
Genetic code is degenerate (each AA encode by more than one codon).
10
Q
What is the primary structure of a protein?
A
Sequence of a chain of AA.
11
Q
What is the secondary structure of a protein?
A
Hydrogen bonding of the peptide backbone causes the AA to fold into repeating pattern.
12
Q
What is the tertiary structure of a protein?
A
3D folding patterns of a protein due to side chain interactions.
13
Q
What is the Quaternary structure of a protein?
A
Protein consisting of more than one AA chain.
14
Q
What does translation involve?
A
involves to specialised types of RNA.
15
Q
What is rRNA?
A
Ribosomal RNA
Forms a molecular machine that conducts translation.
Large and small subunits
16
Q
What is tRNA?
A
Transfer RNA.
Carries specific AA into the ribosome during translation
Has a feature called an anticodon.
17
Q
Where is translation performed?
A
Ribosome
18
Q
What does a ribosome have?
A
has a small and a large subunit
Each subunit is made of proteins and ribosomal RNA (rRNA)
19
Q
What does a 1/3 of the ribosome contain?
A
1/3 of mass of the ribosome is protein
Proteins on the exterior
20
Q
What do rRNA catalyse?
A
rRNA molecules catalyse the formation of the peptide bond between amino acids
21
Q
Two of the three domains populated exclusively by ______.
A
prokaryotes
22
Q
In the third domain _____ multicellular life is largely restricted to ____ recent branches ____, ____, _____.
A
Blank 1-(Eukarya)
blank 2-three
Blank 3- (fungi, plants, animals)
23
Q
What does the anti-codon correspond with?
A
The codon in order to form AA.
24
Q
What do all proteins start with?
A
the amino acid Methyonine (M)
25
What do the subunits do when attached to an mRNA molecule?
The small and large ribosomal subunits assemble when attached to an mRNA molecule
Translation initiation complex
26
What is the P-site?
peptidyl site), binds to the tRNA holding the growing polypeptide chain of amino acids.
27
What is the A site?
(acceptor site), binds to tRNA holding the new amino acid to be added to the polypeptide chain.
28
What is the E site?
exit) binds a tRNA that already unloaded its amino acid and it is going to be released.
29
What is termination?
A release factor binds directly to the stop codon in the A site
Addition of H2O molecule instead of an amino acid
30
What is the process of termination?
1) Ribosome reaches a stop codon on mRNA.
2) Release factor promotes hydrolysis.
3) Ribosomal subunits and other components dissociate.
31
In Prokaryotes when is mRNA translated into proteins?
In prokaryotes mRNA is translated into protein as soon as it is made
Because transcription and translation happen in the same place.
32
How is mRNA translated into proteins in eukaryotes?
In eukaryotes mRNA has to be transported out of the nucleus before it can be translated
33
What do both transcription and translation have in common in eukaryotes and prokaryotes?
In both cases several ribosomes can simultaneously translate one mRNA molecule
34
What is a consequence of having a genetic code?
Reading frames
35
What is a open reading frame?
Start with the AUG start codon
| End with one stop codon, UAG, UAA or UGA (U in mRNA, T in DNA).
36
What are operons in bacteria?
Region of DNA containing a cluster of genes under the control of a single promoter
37
How can the genes within the operon be activated?
All the genes of the operon can be activated with a single transcription factor
38
What is the lac operon?
Bacterial operon to metabolise lactose (sugar)
Repressed if there is glucose
Active if there is lactose
39
What happens when there is no lactose present?
When lactose is absent the lac repressor binds tightly to the operator. It gets in RNA polymerase's way, preventing transcription.
40
What happens when there is lactose present?
Rearranged lactose binds to the lac repressor and makes it let go of the operatoe. RNA polymerase can now transcribe the operon
41
What do prokaryotes organise?
organise genes into operons- unit containing several genes involved in the same metabolic pathway.
42
What do eukaryotes typically control?
Eukaryotes typically control expression of individual genes rather than operons like prokaryotes
43
What are there many levels of?
Many levels of regulation!!! From chromatin packing to post-translational modifications of the end protein
44
what are the multiple scales of regulation of gene expression?
```
Epigenetic
Transcriptional
Post-transcriptional
Translational
Post-translational
```
45
What is Transcriptional?
Modify the rate of gene transcription using transcriptional enhances upstream of the promoter.
46
what is meant by Post-transcriptional?
Stability of mRNA, alternative splicing.
47
what is meant by Translational?
speed of translation
48
what is meant by Post-translational?
modification of protein
49
What do transcription factors recognise?
recognize and bind to double stranded DNA
50
What can transcription factors be?
They can be activators or repressors
51
What do transcription factors do?
will bind to DNA which will signal the RNA polymerase to start transcription.
52
What do individual transcription factors do?
regulate the expression of many genes
53
Why are Transcriptional cascades important?
are important for signal transduction and environmental adaptation
54
Outline the steps of the transcription cascade.
Distribution of information within cells and genomes
Sequencing of several steps of gene expression over time
Amplification of a weak signal from a cell surface receptor
55
What do Transcriptional signalling cascades allow?
allow specificity of stimulus-response coupling.
| Environmental information has become ‘encoded’ by these signalling pathways
56
what is gene expression?
The overall process of producing a protein from the gene that encodes the protein
57
What are all aspects of the gene expression process?
Promoter activation
Initiation of transcription and translation
Alternative splicing
Translation
Protein folding and targeting (to parts of the cell)
58
Gene expression is not just a process, it is _____.
quantifiable
59
Gene expression is the whole process of ___ ___ ____.
DNA->mRNA->protein
60
Cells or organisms contain ________ of different types of ______.
The whole ________.
thousands.
mRNAs
transcriptome!
61
What is a transcriptome?
collection of all the mRNAs present in a cell in a given time.
62
What does Northern blotting measure?
mRNA
63
What does Southern blotting measure?
DNA
64
What does Western blotting measure?
Protein
65
what is the use of the Hybridization probe in Northern blotting?
DNA or RNA fragment, labelled with radioactivity or fluorescence. Probe sequence is complementary to gene(s) of interest in the sample.
66
Outline Northern blotting
1)RNA isolated from the organism is separated on a gel (RNA gel electrophoresis)
2)The RNA within the gel is blotted onto a “membrane” that binds to RNA
3)A short piece of single-stranded DNA, complementary to the RNA of interest, is incubated with the membrane
4)The DNA probe only anneals to the mRNA of interest on the membrane.
The DNA probe is radioactive or fluorescent, so detectable
67
What is the gel made up of in northern blotting?
A gel of an aequeous polymer
| usually agarose
68
What do the pores in the gel provide?
The pores in the gel act as a sieve, so smaller mRNA molecules travel faster and further
69
The ____ ____ the mRNA, the _____ _____sticks to it.
Blank 1- more abundant
| Blank 2- more probe
70
How is the amount of probe measured?
measured imaging instruments
71
What does the band thickness tell you?
(band thickness tells you expression level)
| Expression of mRNA associated with insect eye colour, at different developmental stages
72
What is RT- retrotranscription?
A biological function where you need to copy the RNA genes in the genome in the host in order to make copies.
73
what is Retrotranscriptase?
viral enzyme that synthesizes ssDNA from mRNA (copy DNA or cDNA).
74
What does RT synthesise?
RT synthesizes cDNA of ALL the genes in the tube
75
What does PolyT primer bind to?
PolyT primer binds to mRNA poliA tail.
76
What occurs during RT-PCR?
Now we select gene of interest with primers and PCR
The quantity mRNA from each gene determines the quantity of cDNA copied from the mRNA.
PCR using primers
specific to gene of interest
77
What does the band intensity tell you?
Band intensity tells you expression level
| more mRNA for the gene -> more cDNA -> more amplified DNA on the gel
78
What are the limitations of RT-PCR?
used to monitor gene expression over time but is an approximate so we may need something more accurate.
79
What does RT-PCR with qPCR?
A modern elaboration of RT-PCR removes the need to run gels:
DNA-binding fluorescence is mixed with the RT-PCR reaction
The amount of fluorescence is measured during the PCR by a laser
Quantitative PCR or qPCR
80
What do microarrays allow?
allow the study of whole transcriptomes
81
Outline what microarrays allow?
Array of tens of thousands of cells
Each well/cell in the grid contains unique short RNA or DNA probes complementary to the genes of interest
Array is incubated with mRNA or cDNA of a sample
Fluorescent labelling allows detection of the amount of nucleic acid bound to each short DNA sequence on the array
82
what do microarrays require?
specialized scanning and analytical approaches
83
What will the microarrays show?
Spot intensity provides mRNA quantity for each single mRNA
Each cell has one probe
more molecules in the sample more probes will bind to the genes
84
What does RNA sequencing allow?
also allows study of whole transcriptomes
85
What does the next generation sequencing allow?
allows whole genomes to be sequenced relatively cheaply
86
The number of times a ____ ____ ____ for the _____, from each gene, reflects the mRNA ______ level
Blank 1- sequence is obtained
Blank 2- mRNA
Blank 3-expression
87
Outline the process of Measuring the amount of a protein: Western Blotting.
1) The mix of proteins extracted from an organism is separated using polyacrylamide gel electrophoresis (PAGE)
2) PAGE gels are very thin and normally arranged vertically
3) The protein extract is treated with a detergent that gives it a negative charge
88
Proteins have different chemical properties so we have to use __________ instead of agrose.
polyacrylamide
89
what do darker bands indicate in western blotting?
Darker bands indicate that more antibody has attached, so more of the protein of interest is present
90
Why do we need to get all the proteins to have the same negative charge?
After we apply an electric current it will cause protein to travel down to the positive side.
91
_____ _______ studies cross all scales of biology.
Gene expression
92
What provides insights into the expression of all the genes in an organism?
Microarrays and RNA sequencing.
93
What is a mutation?
Mutations are changes in the genome of an organism, also called genetic polymorphisms at population level.
94
Why are mutations essential?
essential for the process of evolution
They create genetic variation
A few mutations are advantageous
95
What is a characteristic of most mutations?
Most mutations are deleterious or neutral
96
What is Polymorphisms?
Specific nucleotide has many forms and shapes. Most mutations are going to break the gene.
97
Why do sperm cells have more mutations?
Only mutations in the X chromosome of the sperm cell will get passed on. Sperm cells have more mutations because there are more of them and they are made faster. Eggs only go through one cell cycle, thus there is less duplication.
98
Only mutations in____ _____ _____ pass to the next generation, producing individuals carrying the mutation in all cells
germ line cells
99
How many mutations does the sperm cell and egg cell contribute to?
A human baby has ~140 new mutations when compared to the parents
Sperm contributes with ~130 mutations
Eggs with ~10.
100
What are large scale mutations?
Alteration of chromosome number
```
Alteration of chromosome structure
Deletion
Duplication
Inversion
Translocation
```
101
What are small scale mutations?
Substitutions (1 nt)
Silent mutation
Missense mutation
Nonsense mutation
Insertions or deletions (‘indels’)
Frameshift missense
Frameshift nonsense
Gain/Loss of amino acids
102
Why are substitutions mutations silent?
as changes the AA not the phenotype.
103
What are the point mutations of substitutions?
Silent
Nonsense
Missense
104
What is Missense?
you don’t have a stop codon so the folding of the protein will be a little different.
105
What is a Single sense mutation in sickle-cell anemia?
the hemoglobin will have a different shape and that will effect the amount of O2 uptake
106
What is a frameshift?
A frameshift is where addition or removal of base(s) change the reading frame
107
What may insertions or deletions result in?
There will be either an extra nucleotide or you will loose one. If you loose or add one you are changing the order of the sequence.
AA will move one position to the left, the second half of the protein will look nothing like the original strand.
108
What is Thalassaemias?
mutation which affects the production of the α or β chains of haemoglobin, leading to abnormal ratios
109
What is α-thalassaemia?
synthesis of the α chain absent or reduced – usually as a result of deletion of one or more α-globin genes
110
What is β-thalassaemia?
synthesis of β chain absent or reduced – usually as a result of defective processing of β-globin mRNA
111
What can also lead to phenotypic changes?
Mutations outside coding regions can also lead to phenotypic changes if they alter gene expression
Eg. Mutation in promoter reduces level of transcription
112
What do large scale mutations cause?
Large scale are changing chromosome numbers.
113
What is polyploidy?
Many copies
114
What do Accidents in meiosis or cross-breeding cause?
closely related species can cause duplications in the number chromosomes.
115
How does Polyploidy drive evolution and speciation?
because genes on two sets of chromosomes can assume different functions.
116
What does Whole Genome Duplications (WGD): Vertebrates cause?
This can turn a gemome with 8 chromosomes into 16 and can cause an increased rate of mutation.
117
What is Inversion?
genes cannot not recognize other genes.
118
What is Duplication?
causes the promoter to change so the cell is dividing faster and faster.
119
The cell continuously ______ and _____ its genetic material
Blank 1- monitors
| Blank 2- repairs
120
How is DNA proofread and repaired?
DNA polymerases proofread newly made DNA, replacing any incorrect nucleotides
121
What happens in mismatch repair?
In mismatch repair of DNA, repair enzymes (not DNA polymerase) correct errors in base pairing
122
What happens in nucleotide excision repair?
a nuclease cuts out and replaces damaged stretches of DNA
123
What can repair mechanisms do?
Repair mechanisms can correct mispairing, but must be able to recognise which strand is wrong
124
What may mutations in somatic cells cause?
Mutations in somatic cells may give rise to cancer or mosaicism
125
What is meant by Gynandromorphs?
Mosaics of different sexes.
| Mostly arthropods and birds
126
What are the causes of mutations?
```
DNA replication mistakes
Usually corrected by editing or proof-reading function of the DNA polymerase (3’ to 5’ direction).
Recombination mistakes
Meiosis mistakes
Transposons
Mutagens
Chemicals
Radiation
```
127
____ is damaging to DNA.
Radiation
128
What may radiation do to the DNA?
Radiation may cause breaks in DNA
| UV radiation – creates thymine dimers in DNA
129
What chemical may also cause mutations?
Smoking a pack a day for a year causes 150 mutations in lung cells
130
The genetic code is:
The set of instructions written in the genome
The DNA
The correspondence 3 nucleotides to one amino acid
131
The genetic code is consequence of the structure of…
transfer RNAs (tRNAs)
132
The function of a protein is determined by…
```
Primary structure (sequence of aminoacids)
Secondary structure (e.g. sheets and helix)
Tertiary structure
```
133
The technique using a DNA probe to detect a DNA sequence in a membrane is called
southern
134
The technique using a DNA or RNA probe to detect an RNA sequence in a membrane is called
Northern
135
The technique using an antibody to detect a protein in a membrane is called
western
