Section 2: Cellular and Molecular Biology Flashcards
1
Q
Characteristics common to all cells
A
Arise from pre-existing cells
Genetic info stored as DNA in chromosomes
Proteins synthesised on ribosomes
Selectively permeable plasma membrane encloses every cell
Sub-cellular components suspended in cytosol
2
Q
What is cytosol
A
A semi-fluid substance
3
Q
Cell size range
A
Vary a lot because they exist in diff environments and have diff functions
10-100µm
4
Q
What is angstrom used to measure
A
Size of molecule
5
Q
What can you see with a light microscope
A
Most plant and animal cells
Some larger organelles and bacteria
6
Q
What can you see using super-resolution microscopy or electron microscopy
A
Smaller components, e.g. smallest bacteria, viruses, ribosomes
7
Q
Important parameters in microscopy
A
Magnification (enlargement) of image
Resolution - measure of clarity of image
Contrast - difference in brightness between light and dark areas of an image
8
Q
What is resolution
A
The shortest distance between 2 separate objects that can still be distinguished from one another
9
Q
Types of microscopes
A
Light microscope (LM): - fluorescence microscope
Electron microscope (EM):
- transmission EM (TEM)
- scanning EM (SEM)
10
Q
What is light microscopy used for
A
Used to visualise whole cells and large subcellular organelles (nucleus, chromosomes)
11
Q
How do light microscopes work
A
At bottom, there is a light source which is focused onto a specimen on the stage
Image of specimen is magnified by lenses which can then be projected into the eye or a digital camera
12
Q
Types of light microscopy
A
Brightfield
Brightfield (stained specimen)
Phase-contrast
Fluorescence
13
Q
Light microscopy: Brightfield
A
Stained cells to make them more obvious
14
Q
Light microscopy: Phase-contrast
A
Increases contrast to see cells relative to background
15
Q
Light microscopy: Fluorescence microscope - steps
A
Take specimen and add antibodies, each of which is bound to a diff fluorophore and are capable to binding to and recognising distinct molecules in a cell
Incubate for a period and wash away antibodies that haven’t bound
Put specimen under microscope to study
16
Q
Light microscopy: Fluorescence microscope - what do you see
A
An image where diff fluorophores have bound to diff cells / regions within cells –> indicates cells with diff regions have specific molecules –> differentiate types of cells
17
Q
Light microscopy - advantage
A
Can visualise dynamic processes (not just static)
18
Q
Electron microscopy - disadvantage
A
Can’t watch dynamic processes because have to fix and coat samples (i.e. must be dead)
19
Q
Electron microscopy - how does it work
A
Instead of light, EMs use electromagnets to focus a beam of e- through the specimen (TEM) or onto its surface (SEM)
20
Q
Electron microscopy: TEM - how does it work
A
Take sample and stain with a heavy metal, which binds to certain regions of cell
Specimen is put into TEM and e- are focused down onto specimen
21
Q
Electron microscopy: TEM - what do you see
A
Places where there are heavy metals, the e- can’t get through, but regions where there aren’t many heavy metals, e- will go through
This info about the e- that pass though is collected and translated into an image
Can see lots of internal structures within cell
22
Q
Electron microscopy: SEM - how does it work
A
Coat specimen with a layer of gold
Focus beam of e- which excite secondary e- on surface of tissue of cell
Info from secondary e- being excited is translated into SEM
23
Q
Electron microscopy: TEM vs SEM
A
TEM:
- Resolution 2nm (very good)
- Used to study internal cell structure
- Focus beam of e- through specimen
SEM:
- Resolution 10nm (still pretty good)
- Used to study cell surface and generate 3D images
- Focus beam of e- onto surface of specimen
24
Q
What does cell fractionation isolate cell components based on
A
Size and density
25
Cell fractionation - order of isolation?
Isolates largest organelles first and smallest organelles last via homogenisation
26
Cell fractionation - steps
Centrifuged at 1000g for 10 min - pellet rich in nuclei and cellular debris
Supernatant poured into next tube, centrifuged at 20,000g for 20 min - pellet rich in mitochondria (and chloroplast)
Supernatant poured into next tube, centrifuged at 80,000g for 60 min - pellet rich in microsomes
Supernatant poured into next tube, centrifuged at 150,000g for 3 hours - pellet rich in ribosomes
Isolated organelles taken to study
27
Cell fractionation involves...
Differential centrifugation
28
Prokaryotic cells - structure
No nucleus
Little or no internal structure/organelles
Flagella anchored to cell wall/membrane
29
Prokaryotic cells - flagella
Anchored to a motor through hook
Motor spans over cell wall and plasma membrane
As rotor rotates, flagella rotates --> propels cell along
(cilia also involved in helping cells move but don't have a hook and motor structure)
30
Prokaryotic cells - nucleoid
DNA concentrated here but not enclosed by membrane
31
Prokaryotic cells - ribosomes
Complexes that synthesize proteins
32
Prokaryotic cells - plasma membrane
Encloses cytoplasm
33
Prokaryotic cells - cell wall
Rigid structure
34
Prokaryotic cells - glycocalyx
Outer coating consisting of a capsule or slime layer
35
Prokaryotic cells - fimbriae
Attachment to other bacteria
36
Eukaryotic cells - nucleus
Have a membrane bound nucleus which contains *most* of cell's DNA
37
What structures are present in plant cells but not in animal cells
Cellulose cell wall - protects cell and maintains shape
Central vacuole - storage and breakdown of waste products
Chloroplasts - photosynthetic organelle (also present in eukaryotic algae)
38
Organelles / components of eukaryotic cells
```
Endomembrane system
Mitochondria
Chloroplasts (plastids)
Cytoskeleton
Cilia and flagella
Plasma membrane (PM)
```
39
Endomembrane system
Nucleus
ER
Golgi apparatus
Lysosomes
40
Nuclear envelope
```
2 layers (inner and outer membrane) that enclose nucleus
Fuse tgt to form pores within the nuclear envelope
Lined with proteins - molecules can move into and out of nucleus
```
41
Nucleus - what is chromatin
Found within nucleus and is the DNA with histone proteins
42
Nucleus - chromatin structure
When cell is between cell division, chromatin forms a wispy white structure
When cell is about to divide, chromatin is organised into chromosomes
43
Nucleus - nucleolus
Dense region in middle of nucleus
Usually only seen when between cell division
Involved in synthesis of rRNA
44
Outer membrane of nuclear envelope is continuous with...
Membrane of ER
45
What is the endoplasmic reticulum (ER)
An interconnecting network of membranes in the cytoplasm
46
Rough ER (RER)
Ribosomes attached
47
What are ribosomes
Protein-making complexes
48
What are ribosomes composed of
A large subunit and a small subunit
49
Where are ribosomes found
On RER or free in cytosol
50
As ribosomes on surface of RER make proteins...
They insert them into inside of RER
| These proteins are destined to either be secreted by cell or used by membrane-enclosed organelles within the cell
51
Smooth ER - structure
More tubular than RER
52
Smooth ER - function
Involved in synthesis and transport of lipids
53
Relationships among organelles of endomembrane system
Proteins inserted into RER move toward outer layers ER and bud off into a vesicle
These proteins aren't complete and need to be modified --> taken from RER to Golgi apparatus
54
Golgi - structure
A flattened stack of membranous sacs
55
Golgi apparatus - process/steps
Membrane of vesicle fuses with membrane of Golgi, emptying proteins into Golgi apparatus
Proteins can now be modified in several ways - addition of molecules (e.g. carbohydrates, phosphorylated, lipids) that enable them to carry out their functions
Once protein is completed, it's sorted into a particular area of Golgi so it can bud off into a new vesicle
Finished protein can now leave Golgi apparatus
56
Golgi apparatus - where do vesicles fuse
Always fuse at cis side of Golgi, and as proteins move through Golgi, they move to the trans face
57
Golgi apparatus - finished protein - destined to be?
Destined to either by secreted by cell - vesicle buds off and vesicle membrane fuses with outer PM and contents are secreted to outside of cell
OR may be used by other members of endomembrane system - proteins trapped and trafficked back to ER or to earlier regions of Golgi where they may be required to facilitate modifications of proteins
58
What are found in lysosomes
Hydrolytic enzymes made by RER and modified in Golgi, which were then budded off into a lysosome
Hydrolytic enzymes must be enclosed within lysosomes to keep them away from and protect other areas of cell
59
Lysosomes - functions
Digestion of food
| Recycling molecules
60
Lysosomes - digestion of food
Food is taken up into a food vacuole by cell
Membrane of lysosome fuses with membrane of food vacuole and empties its hydrolytic enzymes into food vacuole --> digests food into molecules that can be used by cell
61
Lysosomes - recycling molecules
Enables cells to break down damaged organelles by emptying its hydrolytic enzymes into vesicle --> liberates the molecules which are in those organelles --> enables cell to use and recycle those molecules
62
What are mitochondria
Respiratory enzymes (Kreb's cycle) located in inner membrane and matrix
63
What do mitochondria contain
Own mitochondrial DNA and free ribosomes - potential to produce own proteins
64
Mitochondria - membranes
2 membranous structures - outer and inner membrane
65
Mitochondria - inner membrane
Folds in and out of mitochondria to form cristae
66
Mitochondria - what occurs here
Site of cellular respiration and where oxygen and food molecules are combined to make ATP for cell
67
Mitochondria - importance of cristae
Important because lots of enzymes involved in respiration processes are embedded in cristae
Folds = increased SA = increased no of enzymes present in mitochondria
68
Mitochondria - where are enzymes found
Not all enzymes embedded in cristae, some found in matrix (region between cristae)
69
Where are chloroplasts found
Only in plants and algae
70
Chloroplasts - what occurs here
Photosynthesis
71
Chloroplasts - membranes
Bound by an outer and inner membrane
72
Chloroplasts - thylakoids
A 3rd internal membrane network containing photosynthetic apparatus
73
What do chloroplasts contain
Own DNA and ribosomes
74
Chloroplasts belong to a family called ____
Plastids
75
What are thylakoids
Interconnected flattened sacs stacked on top of each other
| Contains chlorophyll
76
Cytoskeleton - functions
Maintain cell shape (all)
Facilitate cell movement
Facilitate movement of components within cell (e.g. vesicles)
77
What is the cytoskeleton
Interconnecting protein structures within cytoplasm
78
Types of cytoskeletal filaments
Microtubules
Microfilaments
Intermediate filaments
79
Cytoskeleton: Microfilaments (actin)
Cell motility - enable formation of pseudopodia (cell 'foot') and to extend out and crawl along
80
Cytoskeleton: Intermediate filaments
Involved in helping anchor organelles in position inside cell
81
Cytoskeleton: Microtubules
Provides a network for vesicles to move along using microproteins attached to bottom of vesicle
ATP enables motor protein to change in shape --> walks along microtubule, taking with it the vesicle containing the protein to Golgi to be modified
82
Cilia and flagellae contain...
Microtubules
| Help flagella bend and cilia waft
83
Cilia and flagellae - similarity of structure
Similar internal structure
84
Cilia and flagellae - microtubules
9 pairs around the edge and 1 pair in the middle
Outer microtubules linked by motor proteins called dimes
Allows it to bend to an extent (using ATP), then goes back because of cross-linking proteins in between
85
Cell membrane - function
Forms a barrier that selectively regulates movement into and out of cell (PM)
Also membranes surrounding organelles - have similar structure to outer PM
86
What cells have membrane-bound organelles
Eukaryotic cells
87
Lipid bilayer thickness
7-8nm thick
88
Phospholipid bilayer - structure
Each layer contains lots of phospholipids
| Each phospholipid consists of a hydrophilic head and 2 hydrophobic tails
89
Phospholipid bilayer: Head - structure
```
From top to bottom:
Choline (or other small molecule)
Phosphate
Glycerol
(then tail)
```
90
Phospholipid bilayer: What are the tails attached to
2 Cs of glycerol molecule
91
Phospholipid bilayer: Tail
Hydrophobic
| Fatty acid chain
92
Phospholipid bilayer: Tail - saturation
If structure is saturated --> straight
| If structure is unsaturated (has double bond between 2Cs) --> kink in tail
93
For proteins to be embedded in the membrane...
They must have both hydrophobic and hydrophilic regions
Hydrophobic regions to associate with fatty acid tails in middle of membrane
Hydrophilic regions to associate with hydrophilic heads and aqueous environment either inside or outside of cell
94
Amphipathic
Have both hydrophilic and hydrophobic properties
95
Types of proteins in cell membrane
Integral proteins
| Peripheral proteins
96
Integral proteins
Can span entire membrane - called transmembrane proteins
| Or, can only partially protrude into cell membrane
97
Peripheral proteins
Can associate with phospholipids themselves or with integral proteins
98
Glycoproteins
Proteins attached to carbohydrate structures
| Many receptors and docking proteins embedded in cell membranes are glycoproteins
99
Proteins anchored to cytoskeleton
Some proteins embedded in membrane can be anchored to cytoskeleton inside cell
Anchors cytoskeleton in position --> helps maintain cell shape and size
100
In animal cells, instead of having a cell wall like plants, they have...
ECM - a network of fibres outside the matrix
101
ECM components
Proteins and carbohydrates
| Made by the cell themselves (makes its own)
102
ECM - collagen fibres
Main component of ECM
Long strong protein fibres
Intertwined with a proteoglycan complex
103
Proteoglycan complex
Consists of a core protein with numerous carbohydrates radiating off the side
104
ECM - fibronectin
Links proteins embedded in cell membrane with collagen proteins
Can anchor cell in a particular position within the tissue
Enables cell to follow collagen fibres and navigate its way through the tissue
105
ECM components can interact with...
Integrins, which can detect changes in ECM and communicate these changes to inside of cell through a signalling cascade
106
Functions of membrane proteins
```
Enzymatic activity
Signal transduction
Cell-cell recognition
Attachment to cytoskeleton and ECM
Intercellular joining (cell junctions)
Transport
```
107
Functions of membrane proteins: enzymatic activity
e.g. protein enzymes embedded in cristae which carry out respiration
Enzymes are embedded in sequence - enables to catalyse sequential reactions
108
Functions of membrane proteins: signal transduction
Act as receptors to detect signalling molecules in extracellular environment
Once receptor bound to signalling molecule, it can communicate inside cell via signal transduction
109
Functions of membrane proteins: cell-cell recognition
Glycoprotein attached to a small carbohydrate and another protein can recognise this specific glycoprotein --> enables cells to dock with each other briefly --> enables other proteins in membrane to communicate with each other while they are docked tgt
110
Functions of membrane proteins: attachment to cytoskeleton and ECM
Anchors cytoskeletal fibres in place and helps maintain cell structure and place
Or can associate with collagen fibres of ECM
Or may move through tissue by following collagen fibres
111
Phospholipids are synthesised in association with...
The ER
112
Synthesis of membrane components and their orientation - steps
At each transition with ER and Golgi, the blue layer (hydrophobic) of phospholipid remains facing inside of those organelles
Orange layer always faces outer cytoplasm of vesicles and Golgi
As membrane of vesicle fuses with outer PM, orange layer remains facing inside of cytoplasm, but blue layer now faces outside of cell
113
For a protein to be embedded in the membrane and conduct its function correctly...
It must be correctly orientated in the membrane
| Thus, its important when the protein is first made in the ER, it is orientated correctly
114
Where can proteins throughout cell be embedded
Some embedded in inner membrane to carry out their functions, but others need to be embedded in outer PM of cell (e.g. receptors and docking proteins)
115
Fluidity of phospholipids
Phospholipids move very rapidly within their monolayer - lateral movement occurs ~10^7 times per second
Phospholipids rarely flip-flop from one side of PM to the other because hydrophilic head would have to cross hydrophobic tail - ~once per month
116
Fluidity of membrane - double bond
Helps maintain fluidity as it can push away neighbouring phospholipids --> creates more space
117
Fluidity of membrane - if all phospholipids had fully saturated fatty acid tails..
They would pack tightly tgt and end up with a very viscous membrane (not much space for phospholipids to move)
118
Fluidity of membranes - animal and plants living in very cold temp
Have lots of unsaturated fats in phospholipids because otherwise they would stick to each other and solidify - helps retain fluidity of membrane
119
What cells is cholesterol found in
Only found in animal cell membrane - not plants
120
Fluidity of membranes: Cholesterol
Acts as a fluidity buffer (in animal cell membranes)
| A steroid which can squeeze between fatty acids of phospholipids
121
Fluidity of membranes: Cholesterol - temp
At cold temp, stops neighbouring phospholipids from sticking tgt and solidifying - helps maintain fluidity
At moderate temp, reduces amount of space between phospholipids and reduces membrane fluidity
122
Cell fusion - process
Mouse + human cell with membrane proteins joined tgt to form a hybrid cell
Leave for one hour, resulting in mixed proteins
Indicated proteins were able to move laterally around the membrane - membranes aren't static!
123
Cell junctions in animal tissues
Tight junction
Desmosome
Gap junction
124
Cell junctions in plant tissues
Plasmodesmata
125
Cell junctions: Tight junction
Mesh of proteins knitting 2 neighbouring cells tightly tgt --> fluid unable to pass between cells
Primarily composed of occludins and claudins
e.g. skin cells - makes us water-tight
126
Cell junctions: Desmosomes
Anchor is extremely strong between 2 neighbouring cells
Anchored in position by intermediate filaments (keratin) that radiate into cell
e.g. muscle cells
127
Cell junctions: Gap junctions
Each junction made of 6 cylindrical proteins called connexins, which form a tube that runs from one cell to a neighbour cell
Tube is ~2nm in diameter, so small molecules can move from cytoplasm of one cell to cytoplasm of neighbouring cell --> allows communication
Types of molecules that can move through are iron, sugars and amino acids
e.g. found in lots of tissues; heart muscle, liver, embryos
128
Cell junctions: Plasmodesmata
Little tubes that run from one cell to another
Similar to gap junctions; facilitate movement of molecules from one plant cell to another
Through these tubes, small proteins and RNA can move
Cytoplasm of 2 neighbouring plant cells connected --> communication
129
Types of transport for small molecule across cell membrane
Passive transport
- Diffusion
- Facilitated diffusion
- -- Channel proteins
- -- Carrier proteins
Active transport
- Na/K pump
130
Passive membrane transport
Molecules move passively from high to low conc
| Spontaneous - no energy expenditure by cell required
131
Passive membrane transport: Diffusion
No protein required
| Small, non-polar molecules (e.g. CO2 and lipids) are hydrophobic, so can easily diffuse through cell membrane
132
Passive membrane transport: Facilitated diffusion
```
Polar molecules (e.g. water, ions and glucose) are hydrophilic, hence require transport proteins to diffuse through membrane
Most transport proteins are specific --> selectivity
```
133
Osmosis
Diffusion of water across membrane
134
Passive membrane transport: Facilitated diffusion - channel protein
Provides a channel through which specific molecules can move from a high to low conc
e.g. aquaporin - facilitates movement of water from one side of molecule to the other
135
Passive membrane transport: Facilitated diffusion - carrier protein
Highly selective
Has a receptor that enables binding of a specific molecule
When that molecule binds, it induces a change in shape --> molecule has access to other side of cell membrane
As molecule dissociates from protein, protein quickly reverts back to original shape so it can move another molecule
e.g. glucose transporter
136
Active membrane transport
Molecules move across membrane against conc gradient
Usually require energy in form of ATP
All carrier proteins
137
Active membrane transport: Na/K pump - purpose
Allows cell to maintain a low conc of Na+ and high conc of K+ --> cell is able to maintain its resting potential
138
Active membrane transport: Na/K pump - steps
Na/K pump has a carrier protein which has 3 binding sites for Na+
As the 3rd Na+ binds, it enables ATP to donate a phosphate group --> carrier protein now phosphorylated
This induces a change in shape, and 3 Na+ now have access to outside of cell
Simultaneously, the 3 binding sites for Na+ reduce their affinity to bind Na+, so Na+ is quickly released to outside of cell
Phosphate group that was bound to carrier protein dissociates from carrier protein, enabling K+ binding sites to develop a higher affinity for K+
This induces a change in shape and K+ now have access to inside of cell
Simultaneously, affinity of K+ binding sites are reduced to K+ quickly released
Cycle repeats
139
How much ATP made by a cell is used to run the Na/K pump when resting?
~30%
140
Membrane transport proteins - advantage(s) and disadvantage(s)
Provide selectivity
Can increase rate of transport
Continuously recycled
Rate of transport limited by no of proteins
141
Types of transport for large molecules across cell membrane
Exocytosis
Endocytosis
- Phagocytosis - cell 'eating'
- Pinocytosis - cell 'drinking'
- Receptor mediated endocytosis
142
Exocytosis
Where contents of a cell vacuole are released to the exterior through fusion of the vacuole membrane with the cell membrane
143
Endocytosis: Phagocytosis
Cell rearranges to cytoskeleton to send out 2 pseudopodia on either side of the food particle to engulf it into a food vacuole
144
Endocytosis: Pinocytosis
Where cell takes little 'gulps' of ECF containing some molecules - non-selective
As it takes a 'gulp', it pinches the ECF and encloses it within a vesicle
Once vesicle is formed, it is coated with proteins --> coated vesicle
145
Endocytosis: Receptor-mediated endocytosis
Receptors on cell surface recognise and bind to molecules and cluster tgt on membrane --> pinched off into a vesicle
A few other molecules in ECM are also engulfed within the vesicle, i.e. both specific and non-specific uptake
Eventually buds off into a vesicle and is coated with proteins --> enables it to be trafficked to right place of cell
Empty receptor moves back to outer cell membrane
146
Reuse of receptors?
Receptors are recycled - not just used once
147
Functions of proteins
```
Enzymes
Defensive
Storage
Transport
Hormones
Receptors
Contractile/motor proteins
Structural proteins
Gene regulatory proteins
```
148
How much of the dry weight of most cells is protein?
>50%
149
Functions of proteins - enzymes
Majority of enzymes are proteins
| Selective catalysts that can accelerate rate of specific metabolic reactions
150
Functions of proteins - defense
Immune system - protects against disease
151
Functions of proteins - transport
e.g. Haemoglobin - transports O2 from lungs to other tissue in body
152
Functions of proteins - hormones
Majority of hormones in our body are made of proteins
| Hormones coordinate an organism's activity
153
Functions of proteins - receptors
Proteins embedded in cell membrane
| Detect specific molecules outside cell and communicates to inside of cell
154
Functions of proteins - contractile/motor proteins
e.g. flagella - a motor protein is anchored to one pair of the microtubules, and the other side of the motor protein walks along a neighbouring pair of microtubules --> enables flagella to bend
155
Functions of proteins - structural proteins
e.g. cytoskeleton
| Also have outside cell in ECM, e.g. collagen
156
Functions of proteins - gene regulatory proteins
Combine with specific regions of DNA and control whether certain genes will be expressed
Allows cells to respond to changes in environment
Allows cells to differentiate into particular cell types with specialised functions
157
Parts of an amino acid
```
α-carbon
Amino group (NH2)
Carboxyl group (COOH)
H attached to α-carbon
R side chain
```
158
Amino acids - types of side chains
Non-polar: hydrophobic
Polar: hydrophilic
Electrically charged: hydrophilic
159
Polypeptides are built up with...
Monomers called amino acids
160
Amino acid side chains
Each amino acid has a distinct side chain / R group
Determines how the amino acid behaves in polypeptide
Can be simple or complex
161
Number of amino acids
20 diff amino acids with diff side chains
162
Amino acids: Non-polar side chains
Hydrophobic
| 9 types
163
Amino acids: Polar side chains
Hydrophilic
6 types
Cysteine is only weakly polar
164
Amino acids: Electrically charged side chains
```
Hydrophilic
2 acidic (-vely charged)
3 basic (+vely charged)
```
165
Peptide bonds
Link carboxyl group of one amino acid to amino group of next
Dehydration reaction
Form one at a time
Forms a repetitive backbone
166
What do amino acid side chains determine
How final peptide folds and ultimately its 3D structure and final function
167
Proteins: Primary structure
Linear arrangement of amino acids in polypeptide chain
Sequence of amino acid is unique to each protein
Dictates secondary, tertiary, and quaternary structure
168
Primary structure: Polypeptide synthesis
Amino acid sequence specified by sequence of nucleotides in DNA
Info in DNA copied into mRNA (transcription)
Info in mRNA determines amino acid sequence (translation)
169
Proteins: Secondary structure - types
Alpha helix - regions with repeated coiling
Beta-pleated sheet - diff sheets with repeated folding associate with each other
Particular amino acid sequences form alpha helices, while others have a propensity to form beta-pleated sheets
170
Proteins: Secondary structure - H bonds
Form between repetitive regions of polypeptide backbones
Stabilise secondary structre
O in C=O has -ve charge, which is attracted to H in N-H with +ve charge --> H bond
171
Proteins: Secondary structure - H bonds - strength
Individually are relatively weak, but numerous of them form between regions of polypeptide backbone --> relatively strong
172
Proteins: Secondary structure - H bonds - R groups
Formation of H bonds can be influenced by R groups
If R group is v large, can push away neighbouring polypeptides --> interferes with formation of H bonds - known as steric hindrance
Or, R group could carry a charge - interferes with neighbouring H bonds
173
Proteins: Secondary structure - alpha helix
Delicate coil held tgt by H-bonding between every 4th amino acid
Often found in regions of transmembrane proteins that cross the lipid bilayer
174
Proteins: Secondary structure - alpha helix - transmembrane proteins
Each alpha helices region is connected by a non-folded region
Transmembrane proteins often have the C-terminus facing inside the cell and N-terminus facing outside
175
Proteins: Secondary structure - beta-pleated sheet
Two or more strands of same polypeptide chain lay side by side
H bonds form between O and H of neighbouring polypeptide backbones
Make up the core of many globular proteins
Many H bonds = lots of strength
176
Proteins: Tertiary structure
Describes the overall 3D conformation of the polypeptide chain
Stabilised by interactions between side chains
177
Proteins: Tertiary structure - types of weak interaction(s) between side chains
H bonds: between polar side chains
Hydrophobic interactions and vdW: between non-polar side chains
Ionic bonds: between oppositely charged side chains
Individually weak, but tgt strong
178
Proteins: Tertiary structure - types of strong interaction(s) between side chains
Disulphide bridge: type of covalent bond between 2 cysteine amino acids
- Sulfhydryl groups associate with each other, Hs lost and S atoms form a disulphide bridge
179
Proteins: Quaternary structure
The entire protein structure, i.e. all polypeptides
| Consist of two or more polypeptides, which can be the same or diff
180
Proteins: Quaternary structure - Collagen
Consists of 3 identical coiled polypeptides that intertwine with each other
181
Proteins: Quaternary structure - haemoglobin
Consists of 4 polypeptides; 2 identical alpha subunits and 2 beta subunits, which come tgt with 4 haeme groups
182
Chaperonins
Assists folding of proteins (provides optimal environment)
- protects polypeptide from degradation
- polypeptides fold spontaneously into proteins
Some chaperonins tgt with associated systems check correct folding has occured
- refold
- mark for destruction
183
If proteins are misfolded...
It can give rise to various disease states
| Thus important they fold correctly
184
Chaperonins - steps
1. Cap detaches and an unfolded polypeptide enters the cylinder
2. Cap attaches, causing cylinder to change shape --> creates a hydrophilic environment for folding of polypeptide
3. Cap comes off, and the properly folded protein is released
185
Denaturation of a protein
Destruction of the secondary, tertiary and quaternary structure (i.e. 3D shape)
Breakage of H bonds, Ionic bonds, hydrophobic interactions and S-S bonds
186
Causes of denaturation of a protein
Heat - breaks weak bonds (increase in temp --> proteins move further from each other --> bonds break)
pH - changes ionisation patterns of R groups
Reducing agents - reduce S-S bonds to SH
Organic solvents - disturb hydrophobic and hydrophilic interactions
Detergents - disrupt hydrophobic interactions
187
Hydrolysis vs denaturation
NOT the same
| Hydrolysis breaks primary structure, denaturation doesn't affect primary structure
188
Denaturation and renaturation of a protein
In vitro, some denatured proteins can return to their functional shape if denaturing agent is removed and still function
Provides evidence that all info required for a polypeptide to fold into its correct 3D shape is in sequence of side chains
189
Exergonic reactions
Reactants contain more energy than products
Free energy released
Usually catabolic; large molecules --> small molecules + energy
190
Endergonic reactions
Reactants contain less energy than products
Free energy required
Usually anabolic; small molecules + energy --> large molecules
191
Exergonic reactions with enzymes
Ea is lowered
| Change in free energy is unaffected (energy released is same)
192
Activation energy
Energy needed to break/distort bonds to reach transitions state
193
Enzymes
Act as catalysts that lower the Ea, thus increasing rate of rxn
Not consumed by rxn and can be used again
Almost all enzymes are proteins with complex 3D structures which determine its activity (some are RNA)
194
Catalytic cycle of an enzyme - steps
1. Substrates enter active site
2. Substrates are held in active site by weak interactions (usually H and ionic bonds)
3. Substrates are converted to products
4. Products are released
5. Active site available for new substrates
195
Induced fit
Overall shape of enzyme enables it to clasp down on substrate once it binds to amino acids in active site
After being clasped down, additional ionic and H bonds can form
196
The overall shape, size and structure of enzymes ensures...
The amino acid in active sites are able to bind to the substrate and are correctly aligned
197
Catalytic cycle of an enzyme - general equation
Enzyme + substrate(s) -->
Enzyme substrate complex -->
Enzyme + product(s)
198
Enzyme names end in..
-ase
199
Enzyme names: Isomerase
Catalyse intramolecular rearrangements
200
What enzymes hydrolyse RNA and DNA
Ribonuclease
| Deoxyribonuclease
201
What enzymes synthesise RNA and DNA
RNA and DNA polymerase
202
How many reactions can each enzyme typically catalyse in a second
~1000
| Very quickly - each cycle is v brief
203
Enzyme names can indicate...
What the substrate is
204
How are enzymes classified
According to how tightly they bind to the substrate
205
Co-factors
Non-protein helpers for catalytic activity
206
Types of co-factors
Some permanently bound, some bind reversibly with substrate
Prosthetic groups
Coenzymes
207
Co-factors: Prosthetic groups
Organic or inorganic molecules tightly bound to the protein
208
Co-factors: Co-enzymes
Small organic molecules, non-covalently bound to protein
209
Enzyme and substrate conc
Enzyme conc directly proportional to reaction rate (so long as sufficient substrate) - linear relationship
Available enzyme can become saturated with substrate - increases steeply then plateaus at a max
210
How do cells regulate metabolic pathways
By controlling when are where certain enzymes are active
211
How do cells control when/where certain enzymes are active
Switching on and off genes that encode particular enzymes
Regulating enzyme activity once they are made
- conversion of an inactive enzyme precursor to an active form (usually first enzyme made is not active)
- cellular localisation (enzymes may be locked inside organelles in a cell)
- allosteric regulation (inhibition or activation)
212
Allosteric regulation of enzyme activity - oscillation
There is oscillation between the active form and inactive form
213
Allosteric regulation of enzyme activity - active vs inactive form
Active form:
Active sites are open
Inactive form:
Active sites are non-functional / closed off
214
Enzymes - human body
Majority of enzymes in human body have optimal temp of 37°C
215
Enzymes - temp
As temp increases, ROR increases until a max, where the enzymes break and denature (for humans this is 37°C)
216
Trypsin
Involved in breaking down peptides and polypeptides into amino acids
217
Allosteric regulation of enzyme activity - activating an enzyme
If a cell wants an enzyme to be active, it can produce an activator which binds to a site away from the active site and stabilises one of the sub-units within the enzyme in its active state
This active subunit then communicates to other sub-units within enzyme to remain in active states - only need one activator to bind
218
Allosteric regulation of enzyme activity - deactivating an enzyme
If cell wants enzyme to be inactive, it produces an inhibitor which binds to a site away from active site and stabilises one of the sub-units in its inactive form, which communicates to other sub-units to remain inactive
219
Allosteric regulation of enzyme activity - binding of activator and inhibitor
Only binds weakly - can quickly dissociate to enable enzyme to go back to oscillation stage
220
Allosteric regulation of enzyme activity - Regulatory molecules bind to the enzyme via...
Non-covalent interactions
221
Allosteric regulation of enzyme activity: Feedback inhibition
Multi-step pathway, where each reaction is catalysed by a diff enzyme
Metabolic pathway is switched off by the end product (when in excess) binding to and inhibiting an enzyme that acts early in the pathway (usually binds to allosteric regulatory site on first enzyme --> induces change in shape of active site so it's no longer functional)
222
Types of nucleic acids
DNA and RNA - involved in all informational processes in cell
ATP - stores and transports chemical energy within cell
cAMP - involved in intracellular signalling
223
ATP and RNA
ATP is one of the building blocks used to make RNA
224
cAMP is derived from...
ATP
225
Structure of a nucleotide
Phosphate group
Sugar (pentose)
Nitrogenous base
226
Nucleic acids
A polymer of nucleotides
227
How are nucleotides bound tgt
By covalent bonds called phosphodiester bonds - gives rise to a repetitive sugar-phosphate backbone
228
Types of nitrogenous bases
Pyrimidines:
Cytosine (C)
Thymine (T) or Uracil (U)
Purines:
Adenine (A)
Guanine (G)
229
How can nucleic acid structure be stabilised
By H bonds between nitrogenous bases
230
DNA vs RNA - strands
DNA - double-stranded helix
| RNA - usually single strand, but can bend back on themselves - helps form their unique 3D structures
231
What is DNA
The major hereditary material of the cell
232
DNA: Polynucleotide - directionality
From 5' (phosphate group) to 3' (with -OH of sugar)
233
What provides genetic info
Sequence of nitrogenous bases in DNA
234
DNA: 5' vs 3' end
5' is bound to a phosphate group, so no new nucleotides can be added to the 5' end
3' end C is bound to an OH, which is available to bind to a phosphate group of a new nucleotide,
So, can only add a new nucleotide to the 3' end
235
Complementary base pairing
H bonds form between specific base pairs
A-T (or U)
C-G
236
DNA: Anti-parallel
One nucleic acid runs from 5' to 3' in one direction, and other runs from 5' to 3' in opp direction
237
DNA: Nitrogenous bases in middle of helix
Tightly stacked on each other
| Hydrophobic - start to interact with each other through vdW - helps stabilise double helix
238
Chromosomes: Packing of DNA
DNA starts off 2nm in diameter and starts packing with histones
Nucleosome forms
Histone tails radiate out of nucleosomes, which associate with neighbouring DNA strands and nucleosomes --> gives rise to a 30nm fibre
Protein scaffold used to anchor 30nm fibre as it loops round and round to form a 300nm fibre
Chromosomes are highly organised - genes always end up in the same position in the chromatid (700nm)
Not much known about the last step
239
Nucleosomes
Beads on a string
| Where 8 histones (proteins) cluster tgt and DNA molecule wraps around the histone cluster twice --> forms a nucleosome
240
DNA vs RNA - structure
Unlike DNA, structure of RNA molecules is more variable
241
DNA and RNA - sequence of bases
Sequence of bases in DNA specifies the sequence of bases in RNA
242
Types of RNA
Ribosomal (rRNA)
Messenger (mRNA)
Transfer (tRNA)
243
What is rRNA
The RNA component of the ribosome
244
rRNA - function
Helps support structure of ribosome
Involved in protein synthesis
Stable RNA which comprises the bulk of the cellular RNA
245
Where is rRNA synthesised
In nucleolus from highly repetitive DNA
246
What % of ribosomes are made of rRNA
~60% of ribosomes are made of rRNA and 40% made of protein
247
mRNA - function
Conveys info from DNA in nucleus to ribosome, where it specifies the amino acid sequence of the polypeptide
248
mRNA - speed
Synthesised at a fast rate, degraded rapidly, so is present in relatively small amounts
249
Why do we need mRNA
Because DNA can't leave the nucleus, and to get info from DNA to ribosome, need a messenger molecule
250
tRNA - function
Translator - translates nucleotide sequence in mRNA into amino acids during protein synthesis
251
What does a tRNA contain
4 base-paired regions
3 loops including an anticodon
Amino acid attachment site
252
tRNA - anticodon
Anticodon loop binds to codon in mRNA
253
For each of the 20 amino acids, there is at
| least one specific ____
tRNA molecule in the cell
254
Replication of DNA - the basic concept
1. Parent molecule
2. Separation of strands
3. Daughter DNA molecules, each consisting of one parental strand and one new strand
255
Replication - enzymes
Replication is an enzymic process; carried out by a multi-enzyme complex
Must be accurate and fast
256
Proposed models of DNA replication
Conservative model: 2 parental strands re-associate with each other and 2 new strands associate with each other
Semi-conservative model: Each parental strand forms a double helix with a new strand
Dispersive model: Daughter DNA molecules contain a mix of new DNA and parental DNA
257
Which model is DNA replictaion
Semi-conservative
258
Origin of DNA replication in prokaryotes
Each chromosome only has 2 origins of replication - one for each strand
Parental strands are separated and new DNA is made
As replication continues, there are 2 replication forks
In the middle of 2 replication forks is a replication bubble
Replication forks eventually meet and end up with 2 daughter DNA molecules
259
DNA: Prokaryotes - chromosome shape
Circular
260
Replication forks
Where the parental DNA is pulled apart and made available for replication
261
DNA: Eukaryotes - chromosome shape
Linear
262
Origin of DNA replication in eukaryotes
Each chromosome has multiple origins of replication - can have a few hundred or thousand
Occurs in both directions simultaneously
Each bubble has 2 replication forks
As replication continues, replication bubbles fuse and end up with 2 daughter DNA molecules
263
Proteins involved in initiation of replication: Helicase
Enzymes which break H-bonds and untwist double helix at replication fork so DNA strands available for replication
If mutation so helicase doesn't work, no replication fork will be formed
264
Proteins involved in initiation of replication: Single-strand binding proteins
Prevent single strand DNA from re-pairing, bending, or kinking
265
Proteins involved in initiation of replication: Topoisomerase
Breaks, swivels and rejoins parental DNA ahead of replication fork
As DNA unwinds, coils get pushed down, causing region ahead of replication fork to become super coiled --> large amount of pressure on DNA molecule in that region, so topoisomerase relieves strain by breaking sugar-phosphate backbone and unwinding the supercoiled region
266
Proteins involved in initiation of replication: Primase
Synthesises short RNA primers using the parental DNA as a template
267
First bit of nucleic acid made during DNA replication
Is not DNA, but is RNA because DNA polymerase can't add the first nucleotide in a new nucleic acid strand - needs an existing 3' end to add a new nucleotide to
3' end comes from an RNA primer
268
RNA primer - length
Usually quite short - between 5-10 nucleotides in length
269
What is DNA replication driven by
The energy released from the hydrolysis of bonds between phosphate groups in nucleotide being added
New nucleotide has 3 phosphate groups - the 2 phosphate bonds are hydrolysed --> provides energy required
270
DNA polymerase
Catalyses addition of new nucleotide
| Can proofread and repair DNA as its made
271
DNA replication: Leading strand
Continuous synthesis
| DNA polymerase III adds first nucleotide to 3' end
272
DNA replication: Lagging strand
Discontinuous synthesis of fragments in 5' to 3' direction
273
DNA replication: Bi-directional
Occurs in both directions (left and right) simultaneously
274
DNA replication: Anti-parallel
Overall, new DNA strand must be anti-parallel to parental strand
275
DNA replication: Rules
All strands when paired to each other, must be anti-parallel to each other
Can only add a nucleotide to a 3' end
276
DNA replication: Okazaki fragments
Fragments in lagging strand
| Usually between 100-200 nucleotides long in eukaryotic cells
277
DNA replication: Lagging strand - steps
1. Primase generates RNA primer
2. DNA pol III adds nucleotides to primer, forming Okazaki fragment 1
3. Once reaches next RNA primer, DNA pol III detaches
4. Okazaki fragment 2 is primed
5. DNA pol III adds nucleotides, detaching when it reaches fragment 1 primer
6. DNA pol I replaces RNA in primers with DNA
7. DNA ligase forms bond between DNA fragments
8. Lagging strand in region is complete
278
Shortening of ends of DNA molecules
End primer removed, but can't be replaced with DNA as no 3' end available
After 1st round of replication, lagging strand is shorter than template
After 2nd round of replication, new leading and lagging strands are shorter than original template
Ends of chromosomes get progressively shorter
279
What are telomeres
Region of repetitive nucleotide sequence at the end of each chromatid
280
Telomeres - function
Prevent loss of genes near ends
| With each round of replication, gradually get eroded away
281
Telomeres - continuously dividing cells
In continuously dividing cells, telomerase replaces lost telomeric sequences
282
Transcription
The transfer of info from DNA into (pre) mRNA
283
Translation
Nucleotide sequence of mRNA translated into the amino acid sequence of a protein
284
Where does transcription occur
Nucleus
285
Where does translation occur
Cytoplasm
286
What is translation conducted by
Ribosomes
287
Prokaryotic vs eukaryotic cells - RNA processing
Process is similar in prokaryotic cells, but mRNA that is first transcribed can directly be translated
i.e. doesn't need to undergo RNA processing
288
Transcription - basic steps
Initiation
Elongation
Termination
289
Transcription - detailed steps
1. RNA polymerase binds to promoter on DNA
2. DNA strands unwind
3. RNA polymerase initiates RNA synthesis at start point within promoter of template strand
1. RNA polymerase joins complementary RNA nucleotides one by one to 3' end of growing RNA transcript
2. New RNA peels away from template strand
3. Transcribed DNA rewinds into double helix
1. Complete RNA transcript is released
2. RNA polymerase detaches from DNA
290
What is the promoter
A nucleotide sequence that RNA polymerase binds to during transcription
Determines which DNA strand will provide the template for transcription
291
Transcription: Initiation - transcription factors
In prokaryotic cells, RNA polymerase can bind directly to promoter itself
In eukaryotic cells, transcription factors facilitate binding of RNA polymerase to promoter and help initiate process of transcription
292
Transcription: Initiation - unwinding of DNA strands
Usually pulls apart enough of the DNA to expose 10-20 nucleotides at a time
293
Transcription: Elongation - speed
Addition of RNA nucleotides is catalysed by RNA polymerase
| ~40 nucleotides / sec
294
Transcription: Termination - prokaryotes
Proceeds through a terminator sequence in DNA
| Transcribed RNA terminator sequence provides a signal --> polymerase detaches from DNA
295
Transcription: Termination - polyadenylation signal sequence
In eukaryotes
A point in the DNA which is transcribed into the RNA polyadenylation signal, which is bound by specific proteins in the nucleus, which cut the RNA free from the polymerase
296
Average length of mRNA molecule
900-1200 bases long
| Both DNA strands may code for mRNA
297
Genes - overlapping
Some genes may overlap
| e.g. HIV-1 genome consists of 9 genes, some of which are overlapping
298
Transcription: 5' cap and 3' poly-A tail
Only in eukaryotic cells
| Added by enzymes to pre-mRNA
299
Transcription: 5' cap and 3' poly-A tail - functions
Promote export of mRNA from nucleus
Protects mRNA from degradation where it may be exposed to hydrolytic enzymes
5' cap facilitates ribosome attachment
300
Transcription: 5' cap
Added to 5' end
| Consists of a guanine nucleotide that's linked to the pre-mRNA molecule by a triphosphate linkage
301
Transcription: Poly-A-tail
Added to 3' end
| Consists between 50-250 adenine nucleotides
302
Exons
Sequence which will leave the nucleus as mRNA (coding segment) --> translated into amino acid sequences
303
Introns
Non-coding RNA which lies between exons
304
Splicing
Introns are cut out and exons are ligated tgt to form the mature mRNA molecule
305
Where are UTR found
In final mRNA, there are 2 UTR
306
What are UTRs
Untranslated regions
| Not translated into a polypeptide, but info in coding segment is translated into amino acid sequence of polypeptide
307
Introns - functions
Although not part of coding region, will have lots of important functions within cell
e.g. regulate processes of transcription and translation
308
When does splicing occur
After transcription, where eukaryotic cells modify pre-mRNA
309
Splicing: snRNPs
Small nuclear ribonucleoproteins
| Along with other proteins, form a spliceosome on pre-mRNA
310
Splicing: Splice sites
Within the spliceosome
| Where the snRNA pairs with base pairs in the pre-mRNA intron
311
Splicing: Spliceosome
Cuts the pre-mRNA releasing the intron for degradation and splices exons tgt
Catalysed by snRNA (ribozyme)
312
Splicing: Spliceosome releases ____
mRNA
313
What do snRNPs consist of
Proteins and small nuclear RNA, which can pair with splice sites on introns
314
What is a gene
A region of DNA which codes for a functional product, either a polypeptide or an RNA molecule
315
What do exons code for
When spliced tgt and translated into a protein, the original exons code for particular regions of the protein with specific function
316
Codon
A sequence of 3 nucleotides which can be translated into a particular amino acid
317
Some tRNA molecules are bound to..
Amino acids or polypeptides
318
Where are codons and anti-codons located
Codon: mRNA molecule
| Anti-codon: tRNA molecule
319
mRNA specifies...
The amino acid sequence
320
tRNA - structure
A single strand of nucleotides that fold back on itself
4 regions bound to each other by H bonds, formed between complementary base pairs
Has an amino acid attachment site
Anticodon loop to pair with mRNA codon
321
Why is tRNA important
Important for accurate translation to happen
322
Aminoacyl tRNA synthetases
Enzymes that attach the correct amino acid to each tRNA molecule
20 diff synthetases
Gets energy from ATP
323
tRNA is ____ bound to amino acid
Covalently
324
How many tRNA molecules are amino acids bound to
Some only be bound to one, while others can have a few
325
What does every mRNA molecule contain at its 5' end
A ribosomal binding site
326
tRNA binding sites of ribosomes
P site: holds the tRNA carrying the growing polypeptide chain
A site: holds the tRNA carrying the next amino acid to be added
E site: where discharged tRNAs leave the ribosome
327
Translation - basic steps
Initiation
Elongation
Termination
328
Translation - detailed steps (eukaryotes)
Small ribosomal subunit binds to initiator tRNA
5' cap of mRNA binds to small ribosomal subunit
Initiator tRNA scans mRNA molecule for start codon
Anticodon of initiator tRNA H-bonds to start codon
Large ribosomal subunit binds and initiator tRNA is positioned in P-site of ribosome
Translation begins
Anticodon of an aminoacyl tRNA pairs with complementary mRNA codon in A site
rRNA molecule of large ribosomal subunit catalyses formation of peptide bond between new amino acid and growing peptide
Ribosome moves tRNA into neighbouring sites
mRNA moves along with bound tRNAs, bringing next codon into A site
When ribosome reaches stop codon, A site accepts a release factor, which promotes release of polypeptide from tRNA in P site by hydrolysis
Polypeptide released through exit tunnel
mRNA and ribosomal subunits dissociate
329
Translation: Start codon
Always AUG --> initiator tRNA always UAC
| Signifies start of translation
330
Energy for translation
GTP provides energy for all three stages of translation
331
Translation: Release factor
A protein shaped like a tRNA molecule
| Can only bind to a stop codon
332
Translation: Stop codons
UAA
UAG
UGA
333
Signal mechanism for targeting proteins to ER
Signal peptides on newly generated polypeptides can target them to ER
Many polypeptides are modified in the ER or Golgi after they are made, e.g. glycosylation
334
Signal mechanism for targeting proteins to ER - steps
Signal peptides combine with a SRP which combines with its receptor on ER --> enables free ribosome to dock with ER
As ribosome translates info on mRNA into amino acid sequence of polypeptide, that polypeptide can be inserted into the RER
Once translation is complete, ribosome and mRNA dissociate and protein is now inside ER
335
SRP
Signal recognition particle
336
Polyribosome
A single mRNA strand along which many ribosomes are travelling
Each of these ribosomes are synthesising growing polypeptide chains
Enables cell to make proteins quickly
337
Where are polyribosomes found
In both eukaryotic and prokaryotic cells
338
Where does coupled transcription and translation occur
Only in prokaryotic cells
339
Coupled transcription and translation in bacteria
Bacteria can have DNA molecule with RNA polymerase transcribing info in DNA into mRNA
No physical barrier (nuclear membrane) between transcription and in translation and no mRNA processing required
340
Pyrimidines vs purines - structure
```
Pyrimidines = single-ring structures
Purine = double ring structures
```
341
Which carbon of a sugar is the nitrogenous base attached to
1' carbon
342
Which carbon of a sugar is the phosphate group attached to
5' sugar
343
Why is complementary base pairing important for replication of DNA
Ensures the 2 daughter DNA molecules are exact copies of the parental DNA molecule
344
Genetic code - problem and solution
There are only 4 nucleotide bases to specify 20 amino acids
So, the genetic code is a triplet of nucleotides (codon):
Possible combinations = 4^3
345
What is the genetic code
A set of rules that controls how the info is translated from the genetic material (e.g. RNA) into the amino acid sequence
346
Transcription and translation - overall pathway
DNA strand (template) --transcription-->
mRNA --translation-->
Protein
347
Genetic code: Codon table - organisation
Highly organised
348
Genetic code - redundancy
Code is redundant - most amino acids are specified by more than one triplet
So, there is >1 RNA for some amino acids
349
Genetic code - wobble
Some tRNA molecules require accurate base-pairing only at the first 2 positions of the codon and can tolerate a mis-match at the third
350
Genetic code: Start codon
AUG
| Initiates translation but also codes for methionine
351
Genetic code: Stop codons
UAA, UAG, UGA
| Terminate translation - don't code for amino acids, instead bind to a release factor
352
What happens if a start codon is in the middle of a polypeptide sequence
It will just be translated into Met
353
What does 'wild type' mean
Normal
354
Genetic code - universal
Genetic code is shared by most organisms
355
Genetic code: Mutations
Changes in one or a few nucleotides in a sequence can affect protein structure and function (small scale mutations)
Can be a permanent, inheritable alteration in the DNA sequence
May arise spontaneously as a result of low frequency random errors, or induced by external agents which damage DNA
356
Point mutations
Change in a single base pair
357
Types of point mutations
Nucleotide pair substitutions
| Nucleotide pair insertions or deletions
358
Point mutations: Nucleotide pair substitutions
Can result in:
- silent; no effect on amino acid sequence
- missense; an amino acid is changed
- nonsense; creation of premature termination codon - serious
359
Point mutations: Nucleotide pair insertions or deletions
Cause a frameshift, resulting in a missense or nonsense outcome
360
What can point mutations arise from
Mistakes in replication by DNA polymerase
Mutagenic agents, e.g. UV light
Spontaneous chemical reactions in cells
361
Missense - severity
If occurs later in polypeptide, may not significantly influence how the polypeptide folds and protein may still be able to function
But if happens earlier in polypeptide, more likely to alter how the polypeptide folds and change its 3D structure and impair function
But new amino acid may be similar to previous and not have a significant effect on structure/function
362
Nonsense - severity
If occurs toward end of polypeptide, may not have significant effect
If occurs further in, the polypeptide will be incomplete - likely won't function
363
Frameshift
All subsequent codons will also be shifted
364
Point mutation: 3 nucleotide pair deletion
All deleted from same codon
| No frameshift, but one amino acid missing
365
What can DNA be damaged by
Incorrectly paired or altered nucleotides
| Chemical changes of DNA bases, e.g. UV damage can cause a covalent bond to form between 2 neighbouring thymine bases
366
Repairing damaged DNA by excision repair - steps
1. A thymine dimer distorts the DNA molecule
2. A nuclease enzyme cuts the damaged DNA strand at 2 points and the damaged section is removed
3. Repair synthesis by a DNA polymerase fills in the missing nucleotides
4. DNA ligase seals the free end of the new DNA to the old DNA, making the strand complete
367
Gene expression: Prokaryotes
Prokaryote cell gene expression involves response to changes in available nutrients
368
Gene expression: Eukaryotes
Eukaryote cell differentiation is regulated by gene expression
i.e. the genes expressed determine the type of cell it will become
369
Gene expression - totipotent
Many specialised cells don't lose genetic info
| The nucleus is totipotent - theoretically able to program the development of a new embryo
370
Cloning a mammal: The mammary cell is ________
Totipotent
371
Cloning a mammal: What does it show
Nucleus from differentiated mammary cell contains the genetic info required to generate clone of donor animal
372
Cloning a mammal: Dolly - steps
1. Mammary cell donor --> cultured mammary cells
2. Egg cell donor --> nucleus removed
3. Cells fused
4. Grown in culture
5. Implanted in uterus of a third sheep
6. Embryonic development - genetically identical to mammary cell donor
373
Cloning a mammal: Dolly - length of chromosome
Slightly shorter than original mammary cell
374
How much does of their genes does a cell express
Do not express all their genes
Generally only expresses a small % of the possible proteins it can make e.g. a human cell expresses ~20% of its genes at a given time
375
Gene expression - turning on and off
Cells continually turn genes on and off in response to signals from their external and internal environments
376
Gene expression: Prokaryotes - nutrients
Especially sensitive to nutrients in their environment
When they detect a particular nutrient they can metabolise, they make protein enzymes needed to process the nutrient
When that nutrient is no longer there, they stop making those enzymes
377
Regulation of eukaryotic gene expression: Chromatin modification
Genes in tightly packed (condensed) chromatin are usually not transcribed because RNA polymerase can't access the DNA
378
Highly vs less condensed DNA
Highly condensed DNA = heterochromatin
| Less condensed DNA = euchromatin
379
Regulation of eukaryotic gene expression: Chromatin modification - acetylation
Acetylation of histone tails loosen the chromatin structure --> DNA accessible by RNA for transcription
380
Regulation of eukaryotic gene expression: Chromatin modification - methylation
Where methyl groups are added to histone tails --> DNA no longer available to RNA polymerase --> condensation of chromatin and reduced transcription
381
Regulation of eukaryotic gene expression: Epigenetic inheritance
The inheritance of traits transmitted by mechanisms not involving the nucleotide sequence itself
e.g. the genes in condensed regions of DNA which were methylated will be silenced before and after cell division because remain methylated
382
Regulation of eukaryotic gene expression: Initiation of transcription
TFs can bind to specific regions of DNA
Some bind to promoter region and facilitate RNA polymerase binding (transcription initiation complex) - facilitates transcritpion
Some bind to regulatory sequences - can either stimulate or repress transcription
383
What are transcription factors (TF)
Proteins that can initiate and regulate transcription in eukaryotic cells
384
What is the most frequent way gene expression is controlled
Regulating transcription
385
Transcription initiation complex
Where there is RNA polymerase and transcription factors binding, the entire structure is referred to as the transcription initiation complex
386
Regulation of eukaryotic gene expression: RNA processing
Alternative RNA splicing - diff mRNA molecules are produced from the same pre-mRNA
387
Regulation of eukaryotic gene expression: Translation
Can be controlled by regulating factors which mediate initiation of translation
Initiation of translation for some mRNA can be locked by regulatory proteins which bind to UTRs towards the ends of mRNA --> stops mRNA from binding to ribosome --> translation can't start
388
Haemogloblin mRNA - stability
Highly stable
389
What does lifespan of mRNA in cytoplasm determine
How long it can be used for protein synthesis
390
How long does it take for prokaryotic mRNA to degrade
Typically within minutes
391
How long does it take for eukaryotic mRNA to degrade
Can last from hours to weeks
392
Regulation of eukaryotic gene expression: Protein processing and degradation
Protein modification which is required to produce functional proteins is regulated
Protein trafficking to the correct organelle is regulated
Length of time a protein functions regulated by selective degradation
393
Examples of protein modification
Glycosylation
Phosphorylation
If a protein is deemed not required, it these processes will stop --> protein incomplete --> can't conduct final function
394
Regulation of eukaryotic gene expression: Protein processing and degradation - selective degradation
If a protein is no longer required, it can be tagged with a ubiquitin protein which marks it for degradation by proteasomes
395
How can non-coding RNAs control gene expression
By influencing:
- chromatin packing
- translation
- mRNA degradation
396
Examples of non-coding mRNAs
Introns
Micro RNAs
Small interfering RNA molecules
397
Control of gene expression in prokaryotes: Metabolic control
Occurs at 2 levels:
1. Adjust catalytic activities of enzymes already made (rapid response)
2. Adjust production of enzyme molecules by regulating expression of genes encoding enzymes --> none of the enzymes are made (long-term response)
398
What is the most frequent way prokaryotes regulate metabolic processes
Transcriptional control
399
Hydrolysis of lactose
β-galactosidase catalyses the hydrolysis of lactose (disaccharide) into glucose and galactose (monosaccharides)
400
β-galactosidase - low lactose conc
When lactose is absent, only a few molecules of β-galactosidase are present
401
β-galactosidase - high lactose conc
When lactose becomes readily available, the no of β-galactosidase molecules can increase 1000x within 15 mins
402
How can numbers of β-galactosidase increase rapidly
Bacterial mRNA only lasts a few mins, so bacteria can rapidly change pattern of protein synthesis in response to a change in food source
403
Growth curve of bacteria grown on lactose alone
Lag phase: bacteria start to transcribe β-galactosidase mRNA --> translated into β-galactosidase protein
Exponential bacterial growth: bacteria metabolise lactose and grow
Plateaus
404
Lac operon - structural genes
lacZ gene for β-galactosidase
lacY gene for permease
lacA gene for transacetylase
All of which are involved in processing lactose
405
Lac operon - what is an operon
A group of genes coding for proteins with related functions
| Expression of these genes is under the control of a single promoter and operator
406
lac mRNA - negative regulation
A repressor protein binds to the operator to prevent the gene being expressed
407
lac mRNA - positive regulation
A transcription factor or activator bind to promoter and enable RNA polymerase to initiate transcription
408
lac mRNA - negative regulation - when lactose is absent
```
Regulatory gene (lacI) codes for lac repressor protein
lac repressor protein binds to operator and obstructs promoter --> RNA polymerase can't get access
Transcription of lac operon genes can't occur
```
409
Where is lacI located
Next to the operon
410
lac mRNA - negative regulation - when lactose is present
Allolactose (inducer) binds to lac repressor protein, inducing it to change shape (allosteric protein) so it can no longer bind to the operator --> RNA polymerase can bind to promoter
Transcription and translation of lac operon genes can occur
411
What happens when bacteria are grown on glucose and lactose (when both are present)
E. coli preferentially uses glucose first
When glucose is low and plateaus, E.coli will then generate enzymes for lactose breakdown and use lactose as an energy source
412
How is information that glucose is low conveyed to the lac operon
By +ve control of lac operon by activator protein CRP
Low glucose --> cAMP accumulates --> binds to and activates cAMP receptor protein (CRP - allosteric)
Active CRP binds to promoter and facilitates binding of RNA polymerase to promoter --> transcription of lac operon genes
413
What happens when glucose becomes present again (initially just lactose)
Level of cAMP falls so it can no longer activate CRP
CRP detaches from lac operon
RNA polymerase binds less efficiently to promoter
Transcription proceeds at low level, even if lactose is present
414
Lac operon: Mutations in structural genes (Z, Y, A)
May result in production of a non-functional protein
415
Lac operon: Mutations in repressor gene (lacI), operator, or promoter regions
May:
Abolish transcription: e.g. mutation in promoter may destroy RNA polymerase binding site
Permanently turn on transcription: e.g. mutations in lacI may inactivate the repressor (constitutive mutants)
416
If you abolish transcription...
The enzymes will never be made
417
What must occur if a new polypeptide is going to be secreted from a eukaryotic cell
Its signal peptide must target it to the ER
418
Where would you expect to find tight junctions
Between the cells lining an animal's stomach
419
Eukaryotic cilia and flagella bend because of...
A motor protein called dynein
420
A glycosidic linkage corresponds to what protein
Peptide bond
421
If the operator is moved to the end of the lac operon, past all the genes in the operon, what would likely occur?
Genes of lac operon can be transcribed continuously
422
Why is ATP an important molecule for cellular metabolic processes?
It provides energy coupling between exergonic and endergonic reactions
423
A student researcher removes the 5' cap and poly-A tail from an mRNA molecule and inserts this modified mRNA molecule into the cytosol of a eukaryotic cell. What would you expect the student to find?
The mRNA is digested by enzymes in the cytosol
424
What is the most current description of a gene
A DNA sequence that is expressed to form a functional product; either RNA or a polypeptide
425
There is a mutation in the lacI regulatory gene that generates a super-repressor protein, which represses the lac operon permanently. What describes this mutant super-repressor protein?
It cannot bind to the lactose inducer
