Lecture notes from cells module, first year biochemistry. Covers everything you need to know for cell biology, perfect for multiple choice exams and first year biology/biosciences. Got me a first in this module
1 Angstrom = 1 nanometre (nm)
1,000nm = 1 micrometre (um)
1,000,000um = 1 metre
- Bacterial cell = around 0.6um
- Red blood cell = 9um
- Fibroplast = around 50um
- Cells can range from 10nm to 100um
- Below 10nm is chemistry (subcellular) and above is tissues
Robert Hooke (elasticity law) = published ‘Micrographia’ in 1665
- ‘Some physiological descriptions of minute bodies made by magnifying glass
- Reports 60 observations, mainly small things (insects), property of air, small stars +
the moon
- Looked at a thin slice of cork and notices small dots and channels + called then
“pores or cells”. These were actually the cell walls of plants
- Used the word cell similar to a monks cell
Bacteria = flagellum propelling them around
Protozoan = single cell with flagella
Amoeba = single cell, crawls around
Mammalian kidney cell, neuron cell, red blood cells
Fungal = hyphae (many cells) or yeast (single cells)
Anthony van Leeuwenhoek
- 1976 = discovered bacteria
- 1674 = free-living protists
- 1677 = sperm cells
- 1688 = muscle fibres
1839 = ‘Cell theory’ by Matthias Schleiden + Theodor Schwann
1. The cell = unit of structure, physiology and organisation
2. Cells retain dual existence as a distinct entity and a building block
3. Cells form by free-cell formation (similar to crystals)
a. Shown as wrong by Rudolph Virchow
b. 1858 = all cells only arise from pre-existing cells
Francis Crick 1958 = ‘central dogma’ of molecular cell biology
- 3 molecules can interact to pass on detailed info
- DNA to RNA to protein via transcription and translation
Common features of all cells
1. Cells gather material from the environment and duplicate in the cell cycle. This is
divided into M (chromosome inheritance + cell division), G1, S (DNA replication) and
G2
, 2. Information is stored and inherited by DNA
3. Information is partially transcribed into the intermediate form of RNA
4. RNA serves for construction of protein in translation
5. Proteins are the molecules that put genetic information into action
6. All cells are enclosed by a plasma membrane across which materials must pass
Human body = 104 cells
Organic chemistry definition = all cells are basically the same in chemical composition
- All energy flow of life occurs within cells
Five kingdoms of life = plants, fungi, animals, protista and prokaryote (monera)
3 domains of life = bacteria, archae + eukaryote
- In 1977 Carl Woese introduced archaea based on ribosomal RNA sequences
Prokaryotic = cell wall
Eukaryotic = seems more complex with large nucleus + lots of organelles
Monera = no nucleus
Fungi, plants, animals and protozoa are all similar therefore = eukaryotes
- Except animals don’t have a cell wall
Bacteria + archaea = prokaryotes = present before the nucleus + basis of evolution
Microscopy has allowed us to know such detail about the structure of cells
Light microscopy Electron microscopy
Visible light excites/visualises samples Uses electrons (sensitive to air molecules)
Can look at living cells Can only look at dead cells
Glass lens focuses light Electromagnetic lenses
Resolution limit = 20nm Higher resolution limit = 0.05nm
Resolution limit = the minimum distance that allows recognition of object details
- Depends on wavelength of light used
- Smaller wavelength = better resolution
Electron microscopy = always black and white due to the detector (either yes or no electron
detection)
- Scanning electron microscopy can only really visualise surface or outer
- Transmission electron microscopy has higher resolution, light and dark areas can be
seen, can visual inside cells or structires
Advanced electron microscopy
- Single molecule analysis = resolution of 0.001nm (proteins are 10-50nm)
- Freeze fracture electron microscopy = freezes cell causing breakage (tends to be in
between membrane) to see surface of a structure
- Electron microscopy provides ultrastructural information e.g. mitochondria
Multiple electron microscope images are 3D reconstructed using false colours
- This isn’t live cell imaging its called 3D reconstruction
Live cell imaging
- Structure analysis = x-ray electron cryo-microscopy gives detailed structural info
, - Protein-protein interaction studies = fluorescent resonance energy transfer gives
dynamic interactions
- Microarray technology/expression profiling
All this doesn’t explain what’s going on in a cell whereas live cell imaging + the movies give
something extra in combination with other techniques
Principle of fluorescent microscopy:
- Fluorescence is the emission of light by a substance that has absorbed light
- Emission is at a higher wavelength that excitation
- Fluorescence microscope excited the specimen and collects the emission light
- Different fluorescence proteins have different excitation and emission spectra
Green fluorescent protein = GFP
- This is the biggest fluorescent protein seen in Aequorea Victoria jellyfish
- Calcium triggers a protein to give out blue light which is detected by GFP + green
light is sent out
GFP is used to tag and report proteins in living cells by fusing to other proteins (fusion
proteins)
1. GFP + mitochondrial protein gives green mitochondria that can be excited by blue
light
2. GFP + nuclear localisation signal should go into nucleus creating a green nucleus
There are other fluorescent proteins e.g. DsRed
FRAP = fluorescent recovery after photobleaching
- Destroys GFP in certain area + over time is watched to see if it recovers
- In membranes the proteins can be mobile, when fused to GFP the whole membrane
turns green
- High energy light can be used to destroy GFP’s ability to be excited/bleaches it dark
- If the proteins are mobile, more unbleached molecules will mix with bleached
molecules and fluorescent will reappear
- In the nucleus = if part of the nucleus is bleaches there is no recovery
FLIP = fluorescent loss in photobleaching
- Checks behaviour and recovery
- For example can be used to see if proteins in Golgi enter the endoplasmic reticulum
- GFP is present in both Golgi + ER
- GFP is constantly photobleached in the ER
- This means GFP in the Golgi apparatus also goes down as proteins are going to the
ER and are bleached
Photoactivation = enables modified GFP to send out light
- GFP is modified by point mutations leading to different colours
- E.g. T203H mutation = GFP not excited by blue light unless a laser beam is pulsed at
400nm. Refolds GFP that can be excited, 100-fold increase
- Therefore one can activate a subpopulation and see how they interact
, Plasma membrane = specific proteins, lipids + sugars
- Can see in using probes + electron microscopy
- Bilayer
- Made up of amphipathic phospholipids with hydrophilic tail + hydrophobic head
- In the presence of water phospholipids assemble into a lipid bilayer
o This will always form spontaneously due to chemical nature
- Plasma membranes are fluid therefore can be pulled and stretched but hard to
disrupt
- Phospholipids can move and slowly move back
- Same with organelles and vesicles (mitochondria + nuclear envelope)
- Steroids affect membrane fluid but also act as hormones (e.g., cholesterol reduces
membrane fluidity at normal temp but avoids solidification at low temp ‘temp
buffer’)
- Thickness = 4nm
- Overall, biomembranes are fluid with proteins swimming around
Membranes differ in their lipid composition thus, provide some identity to the organelle
- E.g., plasma membrane has little dots with more on the cytoplasmic side than the
exoplasmic side
- These dots are embedded/attached proteins with more on cytoplasmic side as more
is going on
There are specialised lipid-protein regions called ‘lipid rafts’ that assemble specialised lipids
+ proteins to perform certain tasks
- For example, sites of uptake are rich in cholesterol + receptors because this allows
for reduced fluidity, allows uptake and stays together
FRAP shows differences in membrane fluidity + protein mobility
- Photo bleach certain protein in middle so that it cannot be excited
- Free moving molecules mean the area is replaced with excited GFP
- Hence fluid mosaic models
If FRAP is done at the end of a hyphae, the bleaches area doesn’t close suggesting reduced
membrane fluidity + diffusion with is key for growth
Proteins present include:
- Transporters
- Pumps to transport molecules that can’t cross
- Pumps to transport over membranes (cell-cell communication)
- Anchorage + communication to cytoplasm
- Anchorage to extracellular matrix
Types of membrane protein = transporters, enzymes, receptors, cell-cell recognition,
intracellular join + attachment to extra/intracellular matrix or cytoskeleton
- Biomembranes are semi permeable therefore small uncharged or hydrophobic
molecules can diffuse through the membrane
- Charged molecules (sugars, proteins, ions, ATP) cannot pass through
- Channels mediate, communicate + exchange small molecules and ions
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