MGD Flashcards
To cover some aspects of the Molecules, Genes and Disease course for ESA 1 Note: This is not all the content required for MGD, just the shit I struggled with I take no responsibility for any of the flashcards featured here...mistakes/shit happens.
1
Q
What is the name given to diseases caused by improper polypeptide folding?
A
Amyloidoses
2
Q
What type of bonds are involved in the primary structure of proteins?
A
Covalent (peptide) bonds
3
Q
What type of bonds are involved in the secondary structure of proteins?
A
Hydrogen bonds
4
Q
What type of bonds are involved in the tertiary/quaternary structure of proteins?
A
- Hydrogen bonds
- Van Der Waals
- Hydrophobic Interactions
- Covalent (Disulphide)
- Ionic Interactions
5
Q
What are the main features of the alpha helix?
A
- 3.6 amino acids per turn
- 0.54nm pitch
- Right handed helix
6
Q
What are the main features of the beta pleated sheet?
A
- Extended conformation
- Parallel or antiparallel
- Multiple inter-strand H-bonds
7
Q
Which organelles are important for detoxification reactions?
A
- Endoplasmic reticulum
- Golgi body
8
Q
What is the function of the plasma membrane of a eukaryotic cell?
A
- Cell morphology and movement
- Transport of ions and small molecules
9
Q
Give two benefits of a molecule being hydrophobic as opposed to hydrophilic?
A
- Can pass through lipid bilayers
- Can be stored anhydrously
10
Q
What is a zwitterion?
A
A zwitterion is a neutral molecule that has both positive and negative charge
11
Q
When does a zwitterion become deprotonated?
A
When the pH > pKa of an Amino Acid
12
Q
What is the isoelectric point of a protein?
A
The isoelectric point is the pH at which the protein has no overall charge.
13
Q
What two small amino acids are strong helix formers?
A
Alanine and Leucine
14
Q
Why is proline considered to be a helix breaker?
A
The rotation around the N-C bond is impossible.
15
Q
Why is glycine considered to be a helix breaker?
A
The tiny R-group supports other conformations.
16
Q
What effect does 2,3-bisphosphoglycerate (BPG) have on the binding of O2 to haemoglobin? Which way does it shift the oxygen dissociation curve?
A
It decreases the affinity for oxygen binding and shifts the curve right
17
Q
In what two cases is the concentration of BPG increased?
A
- High altitudes
- In highly metabolising tissues
18
Q
What effect do carbon dioxide and protons have on the binding of O2 to haemoglobin? What is this effect called?
A
They decrease the affinity for oxygen binding known as the Bohr effect
19
Q
What type of oxygen binding does myoglobin exhibit?
A
Hyperbolic O2 binding
20
Q
What type of oxygen binding does haemoglobin exhibit?
A
Sigmoidal O2 binding
21
Q
What two states can deoxyhaemoglobin exist in?
A
- Tense state (doesn’t bind oxygen that readily)
- Relaxed state (binds oxygen easily)
22
Q
What is a fibrous protein? Give three examples of what a fibrous protein is used for.
A
One repeating primary structure. Proteins for structure, support, protection
23
Q
What is a globular protein? Give two examples of what a globular protein is used for.
A
Several types of secondary structure structure. Enzymes and regulatory proteins
24
Q
What causes sickle cell anaemia?
A
It’s an autosomal recessive genetic disorder resulting in the substitution of hydrophilic Glutamate to hydrophobic Valine in the β-subunit of Hb
25
What is the pathological progress of sickle cell anaemia?
A ‘sticky hydrophobic pocket’ formed by Val allows deoxygenated Hbs to polymerise, causing sickle shape. This leads to premature destruction of RBCs and blockage of the cell microvasculature.
26
What are Thalassaemias?
A group of genetic disorders where there is an imbalance between α and β subunits.
27
What is Km?
The substrate concentration that gives half the Vmax
28
What is Vmax?
The maximum rate when the enzyme is saturated with substrate
29
List the 5 major regulatory mechanisms that control enzyme activity
- Covalent modification
- Proteolytic activation
- Allosteric control
- Substrate/product concentration
- Enzyme concentration
30
How does proteolytic activation work?
An enzyme secreted as an inactive protein precursor (zymogen) and cleaved by proteases create the active enzyme when needed
31
What type of regulatory mechanism controls phosphofructokinase activity? What activates and inhibits it?
Allosteric regulation:
- Activators are AMP, fructose-2,6-bisphosphate
- Inhibitors are ATP, citrate and H+
32
What is the zymogen of pepsin?
Pepsinogen
33
What is the zymogen of trypsin?
Trypsinogen
34
What is the zymogen of elastase?
Proelastase
35
How many rings does a purine have? What bases are purines?
Two rings (A and G)
36
How many rings does a pyrimidines have? What bases are pyrimidines?
One ring (C, U and T)
37
Which end of the DNA chain has a free phosphate?
5' end
38
Which end of the DNA chain has a free -OH group?
3' end
39
Briefly outline the three main steps of DNA replication.
- Initiation
- Elongation
- Termination
40
What is the direction of chain growth in DNA replication?
5' to 3'
41
What is the genotype?
The genetic make-up of an individual
42
What is the phenotype?
All observable characteristics of an individual or the expressed trait as a result of the genetic make-up
43
Which phase of the cell cycle is all the genetic information duplicated in?
S phase
44
What is the function of the G1 phase of the cell cycle?
Cellular content is duplicated
45
What is the function of the G2 phase of the cell cycle?
Double checking everything is correct
46
What is a gene?
A unit of heredity; a length of DNA on a chromosome that contains the code for a protein
47
What is an allele?
An alternative form of a gene; each individual has two alleles for every gene, which can either be the same or different
48
Briefly describe the process of initiation during transcription
- Recognition at 5'TATA3' box
- Transcription factors bind at this point
- Attract RNA polymerase
49
Briefly describe the process of elongation during transcription
RNA polymerase travels along forming a complimentary RNA strand
50
Briefly describe the process of termination during transcription
- Capping - a methyl-guanine ‘cap’ to the 5’ end
| - Tailing - polyadenylated - lots of Adenine nucleotides are added
51
Outline the process of initiation during translation
- Binding of 40s sub-unit of Met-tRNA to the 5' cap of mRNA
- Codon 5'AUG binds to anticodon 5'CAU
- 60S subunit then binds
52
Outline the process of elongation during translation
- Met-tRNA occupies p site of ribosome and then another aminoacyl-tRNA occupies the A site using GTP
- Formation of peptide bond between amino acids via peptidyl transferase, making the tRNA in P site uncharged (without amino acid)
53
Outline the process of termination during translation
- Stop codon on mRNA reached and no tRNA can bind to it
| - Peptide and tRNA are hydolysed to release the protein into the cytoplasm
54
How can mutations outside the coding region affect gene expression?
Mutations to promoter regions where transcription factors bind can affect gene expression
55
What is regulated secretion?
-Proteins packaged into vesicles but not released until a signal (hormone)
E.g. Insulin
56
What is constitutive secretion?
-Proteins packaged into vesicles and release continuously by exocytosis.
E.g. Albumin
57
Which drug blocks peptidyl transferase?
Chloramphenicol
58
What is the function of the drug tetracycline?
It can block the binding of aminoacyl-tRNA
59
Give the anagram commonly used for collagen synthesis and briefly outline what each letter stand for.
CHADPOGRL:
- Cleavage of signal peptide
- Hydroxylation of proline/lysine residues
- Addition of N-linked oligosaccharides
- Disulphide bond formation
- Procollagen - formation of triple helix
- O-linked glycosylation
- Golgi then exocytosis
- Removal of terminal peptides (procollagen peptidase)
- Lateral aggregations to form fibrils
60
Give a brief overview of the secretory pathway in mammalian cells
- SRP recognises secretory signal sequence at N terminus, binds to sequence and protein synthesis stops
- SRP (attached to sequence) then binds to the SRP receptor on cytosolic face of the ER
- SRP is released, protein synthesis restarts
- Cleavage of signal sequence by signal peptidase after protein synthesis is finished
- Folding of protein, may require chaperones
61
How does protein modification occur in the golgi?
- Trimming and modification of N-linked oligosaccharides
- O-linked glycosylation via glycosyltransferase
- Endo/exoproteases
62
How does protein modification occur in the endoplasmic reticulumn?
- Signal Cleavage (signal peptidase)
- Disulphide bond formation (protein disulphide isomerase)
- N-Linked glycosylation (oligosaccharide-protein transferase and dolichol)
63
Briefly describe O-linked glycosylation of proteins
- Modification of the Hydroxyl Groups on serine/theonine
| - Glycosyl transferase builds up a sugar chain from Nucleotide Sugar Substrates
64
Briefly describe N-linked glycosylation of proteins
- Oligosaccharide is built up on a Dolichol Phosphate carrier molecule in the membrane
- Oligosaccharide is then transferred to the Amide group of Asparagine
65
Where is the "pre" segment of a protein removed?
Endoplasmic reticulum
66
Where is the "pro" segment of a protein removed?
Golgi
67
Outline the formation of the mature insulin molecule
- Removal of signal sequence by signal peptidase (preproinsulin to proinsulin)
- Disulphide bonds form between A and B peptides, cleavage of C peptide by endopeptidases
68
What can C peptide be used as a measure of?
Marker for measuring levels of endogenous Insulin in Diabetics
69
Describe the basic structure of the collagen molecule
- Glycine every third position
- 3 alpha chains
- Left handed triple helix
- Mostly hydroxyproline and proline residues in other positions
70
What is the significance of the proline residues in the collagen molecule?
Prevents the peptide moving into a different structure than the extended α-chain conformation
71
What is the significance of the hydroxyproline residues in the collagen molecule? How are they formed?
Formed from hydroxylation of proline (Prolyl Hydroxylase) and forms many interchain H-bonds
72
What two things are required for Prolyl Hydroxylase to function correctly?
- Vitamin C
| - Fe 2+ ions
73
What is scurvy due to?
Vitamin C deficiency
74
How are proteins targeted to the nucleus from the cytoplasm?
- Fully folded protein with an NLS is bound to the protein importin
- Complex translocates through nuclear pore
- Release of nuclear protein
- Importins are moved back out to be reused
75
What does penicillin do? How does it do it?
It inhibits the formation of bacterial cell walls by inhibiting the transpeptidase enzyme that forms cross-links in cell wall. Which results in osmotic pressure causing cell lysis.
76
What does rifampicin do? How does it do it?
Binds to bacterial RNA Polymerase preventing transcription and therefore preventing DNA replication
77
What does tetracycline do? How does it do it?
Competes with tRNA at A site of bacterial ribosome and therefore inhibiting translation
78
What does methotrexate do? How does it do it?
-Impairs the synthesis of tetrahydrofolate from folic acid - essential for DNA synthesis -Done by competitively inhibiting dihydrofolate reductase (DHFR)
79
Give 5 mechanisms by which cells can become resistant to an antibiotic or drug.
- High rate of division (mutation - chance resistance - natural selection)
- Decreased influx (takes up less of drug)
- Increased efflux (more kicked out by protein channel being upregulated)
- Increased Transcription of Target (increasing transcription of target to overwhelm the drug)
- Altered Target
80
Outline the mechanisms involved in targeting proteins to the mitrochondria (matrix proteins)
- Unfolded and have an amphipathic N-terminal signal sequence
- Stabilised in the cytosol by interaction with molecular chaperones like MSF
- Sequence recognised by TOM proteins in the mitochondrial outer membrane - form a protein import channel
- TIM proteins move across inner mitochondrial membrane (using ATP) and N-terminal signal sequence is removed by MPP
- Protein folds into its native conformation using ATP and chaperones
81
Outline the mechanisms involved in targeting proteins to the lysosome
- Lysosomal Hydrolases are targeted by addition of a Mannose-6-phosphate (M6P) signal to N-linked oligosaccharides
- M6P added by N-acetylglucosamine phosphotransferase/phosphoglycosidase.
- Golgi has M6P receptors to move vesicles containing the protein in the right direction
82
How is I-Cell Disease caused and what does it result in?
- Genetic defects in the N-acetylglucosamine phosphotransferase enzyme result in a lack of M6P addition to lysosomal targeted proteins.
- Lysosomal hydrolases mistargeted for secretion - proteins present in blood/urine
83
How do proteins that are meant to remain in the ER but get released get back into the ER?
- Have a KDEL sequence near C terminus
| - KDEL sequence binds KDEL receptors and is returned to the ER in transport vesicles
84
Briefly outline the Sanger Dideoxy Chain Termination Method of DNA sequencing
- Fluorescent/Radioactively stained ddNTPs (dNTPs lacking a 3’ OH group) are added terminating the sequence at different points
- Produces lots of new DNA fragments of different lengths that can be denatured with heat and separated using gel electrophoresis
85
Briefly outline the process of restriction analysis
- Restriction endonucleases are bacterial enzymes that recognise DNA sequences ‘restriction sites’ and the cut the DNA here
- Cutting of the DNA sequence leaves ‘sticky ends’ - can be reversed/different fragment joined using DNA ligase
86
What can restriction analysis be used for?
- Investigate size of DNA fragments (deletions)
- Investigate mutations (sickle cell disease)
- Investigate DNA variation (DNA fingerprinting)
- Gene cloning
87
Outline the three main steps of gene cloning
- Plasmid is cut using restriction enzymes, gene of interest is added to create ‘recombinant DNA’ molecule
- Transformation - introduced into bacteria
- Multiplication of bacteria
88
Outline the four main steps of gel electrophoresis
- A solution of different fragments is placed in a well at the negative anode end.
- A charge of the anodes encourages the (negatively charged) DNA to move towards the positive anode.
- Larger fragments move slowest.
- Fragments of known size are used as a reference.
89
Briefly outline the process of polymerase chain reaction (PCR)
- Denaturation at high temperature (94 – 96 degrees)
- Renaturation (annealing) at lower temperature (50 - 65 degrees)
- DNA synthesis at medium temperature (75 - 80 degrees)
90
What type of DNA polymerase is used in PCR?
Thermostable Taq DNA Polymerase from Thermus Aquaticus.
91
What is Western blotting an analysis of?
Proteins
92
What is Northern blotting an analysis of?
RNA
93
Briefly outline the process of Southern Blotting and DNA Hybridisation
- Nylon is used to transfer the DNA fragments from electrophoresis
- Hybridised with a labelled gene probe to show the specific DNA fragments
- Radioactive probes can mark specific complimentary DNA fragments.
94
What is the process of Southern Blotting and DNA Hybridisation used for?
- Investigate gene structure (deletions/duplications)
- Investigate gene expansion (triplet repeats)
- Investigate variation (DNA fingerprinting)
95
How can PCR be used in an allele-specific test?
Use primers specific for sequence either side of allele of interest to amplify specific allele.
96
How can restriction analysis be used in an allele-specific test?
- Use restriction enzymes with restriction sites around/within the allele. Analyse the size of fragments produced.
- If the restriction enzyme cuts the wild type but not sample, the restriction site is mutated/missing.
97
How can DNA hybridisation be used in an allele-specific test?
Use a DNA probe that is complementary to either the wild type allele or mutated allele. See which binds.
98
Briefly outline the process of SDS page.
- The detergent SDS (Sodium dodecyl sulphate) denatures protein molecules (breaking 30 structure) and gives the protein a negative charge proportionate to molecular weight.
- Electrophoresis then takes place, with migration from negative to positive electrode, largest molecular weight (more negative charge from SDS) travel further.
99
Briefly outline the process of isoelectric focusing
Proteins are applied to a gel containing a pH gradient - protein migrates until a pH that matches it’s pI – at this point it will have no overall charge and so will stop migrating.
100
Briefly outline the process of 2D page.
Combination of both SDS page and isoelectric focusing - the gel is run in one way and when two proteins stop at an identical value it is run in a 90 degree angle to the original movement
101
What is an enzyme assay?
Methods for measuring enzymatic activity, vital for study of enzyme kinetics/inhibition.
102
What conditions need to be met for an enzyme assay to be taken?
- Optimal pH
- Optimal temperature
- Optimal ionic strength
- Appropriate ions or cofactors
103
How can enzyme assays be used to detect tissue damage?
Elevated levels of enzyme in serum can be indicative of damage to the tissue that usually contains that enzyme
104
Which two enzymes can be used to detect liver damage/disease?
Aspartate transaminase (AST) and Alanine transaminase (ALT)
105
Which two enzymes can be used for detection of a myocardial infarction?
Creatine Kinase (CK) and Lactate dehydrogenase (LDH)
106
Briefly outline the process of Western blotting (don't bother describing SDS page)
Following SDS-PAGE, the separated proteins can be transferred to a nitrocellulose membrane and specific proteins can be visualised by binding with antibodies conjugated to a label
107
Briefly outline the process of Enzyme-Linked Immunoabsorbent Assays (ELISA)
- Primary antibody "stuck" on solid support
- Solution to be assayed is applied to antibody covered support
- Secondary antibody conjugated with an enzyme binds to the antibody-antigen complex
- Amount of binding of second enzyme is measured
108
What would the normal karyotype for a male be?
46, XY
109
What would the normal karyotype for a female be?
46, XX
110
Describe the components of a chromosome
Chromosomes are made up of chromatids which are made up of:
- Histones
- DNA/RNA
- Non histone proteins
111
What is euchromatin?
Lightly packed chromatin often under active transcription (beads on a string)
112
What is heterochromatin?
Tightly packed chromatin (solenoid)
113
What stage of the cell cycle do chromosomes replicate?
S phase
114
What is polyploidy? What is it often caused by?
A numerical change in the ENTIRE chromosome set, multiples of all the chromosomes caused by polyspermy.
115
What is aneuploidy? From this explain the terms monosomy and trisomy.
An abnormal number that is not a multiple of the haploid number:
- Monosomy - a chromosome exists singly without a partner (not in pair)
- Trisomy - A chromosome pair exists as a triplet - one normal and one double
116
How is a ring chromosome formed from one chromosome?
Loss of telomeres or ends of both arms and formation of a ring
117
What is an isochromosome?
Creation of two non identical chromosomes, one is a combination of the two short arms, the other is a combination of the two long arms.
118
What is a Robertsonian translocation?
Q-arms (long) of two acrocentric chromosomes combine to form one ‘super-chromosome’ with the loss of both p-arms.
119
Why is Karyotyping useful?
- Prenatal Screening (raised maternal age, family history)
- Birth Defects (Malformations, Mental retardation)
- Abnormal Sexual Development (Klienfelter’s Syndrome)
- Infertility
120
What is a transversion mutation?
Where a purine is produced instead of a pyrimidine (or vice versa)
121
What is a transition mutation?
Where a different purine is produced than the original purine OR where a different pyrimidine is produced instead of the original pyrimidine
122
Why can point mutations in non-coding regions or outside genes cause problems?
Alter binding sites, promoter sequence, splice sites etc. which can all change how the protein is produced
123
What is a nonsense mutation?
A mutation that change the amino acid specified to a stop codon
124
What is a missense mutation?
A mutation that replaces one amino acid with another
125
What is a silent mutation?
A mutation that does not alter the amino acid specified
126
What is a frameshift mutation?
Addition or subtraction of nucleotides not in multiples of 3
127
Name 4 things that can induce mutations.
- Alkylating agents (Remove a base)
- Acridine agents (Add or remove a base)
- X rays (Break chromosomes/delete few nucleotides)
- UV radiation (creates thymidine dimers)
128
What is the wild type trait?
The trait that is most common in a population
129
What is excision repair?
Damaged DNA is removed by excision of bases and replacement by DNA polymerase.
130
What are the two types of excision repair?
- Nucleotide excision repair replaces up to 30 bases and is used in repair of UV Damage and some carcinogens.
- Base excision repair replaces 1-5 bases and repairs oxidative damage.
131
When does mismatch repair occur and what does it do?
- Occurs when enzymes detect nucleotides that don’t base pair in newly replicated DNA
- Incorrect base paired is excised and replaced
132
What is the response when DNA repair fails?
Protein p53 monitors repair of damaged DNA and if damage is too severe it promotes apoptosis
133
What is formed when a proto-oncogene is mutated?
Oncogene
134
Why is PCR important in the diagnosis of genetic disease?
Most human mutations are single base changes and therefore hard to detect, PCR makes it easier to detect.
135
Name two autosomal dominant conditions.
- Marfan's
| - Familial hypercholesterolaemia
136
Name two X-linked recessive conditions.
- Haemophilla
| - Duchenne muscular dystrophy
137
What type of RNA is most numerous in the body?
Ribosomal RNA (rRNA)
138
What type of RNA has the most kinds?
Messenger RNA (mRNA)
139
Formation of insulin is an example of what kind of secretion?
Regulated
140
Formation of albumin is an example of what kind of secretion?
Constitutive
