MCO test 3 Flashcards
membrane function
semipermeable barrier
detects and interprets changes in extracellular environment
anchorage for proteins and cytoskeleton
lipid structure
diverse
defined by hydrophobicity rather than structure
soluble in organic solvents
phospholipids
amphiphilic/amphiphatic
polar = hydrophillic
fatty acids are hydrophobic
ampiphatic
both properties
membrane typically contains _ lipids
109 lipids
_____ lipid molecules per __ of bilayer
5 x 10^6 per 1 micrometer squared
internal membrane modifications done in ER only found on ___ side
exo
micelle liposome
hydrophobic
fatty acids
terminate with carboxylic acid group
long hydrocarbons (14-24C) 16/18 most common
saturated fatty acids means no
double carbon bond
so straight chain
C:D
number of carbons to numbers of double bonds
C=C
introduces kink in chain causes irregular packing (leading to lower melting point) OR more fluidity
essential fatty acids must be
Obtained through diet
smedley-maclean
Arachnoid acid 20:4
synthesised from clinic acid precursor for eicosanoids
also important for phospholipid bilayer
plays important role in inflammation
(diagram)
phospholipids can be modified by
ester bonds between phosphate groups
common head groups
have biological functions
phosphaldyl choline and phoshatidlynositol can be cleaved inositol and choline are important signalling molecules
PLC is activated process
1)signal transmits to phospholipase (PLC) only some pathways depend on phosphatidylserine other membrane proteins important
2)inositol phosphate is cleaved and transmits signal into cell
sphigomyelin
sphingosine with a fatty acid and hydrocarbon chain andcholien attached
choline
in myelinated axons signal transduction apoptosis
glycolipids
-sugar containing lipids instead of P group
-can be more than one sugar unit
-dereived from sphingosine not glycerol
-sugar always on outside of cell
cholesterol
-modified steroid
-cholesterol 4 hydrocarbons only steroid ring is in membrane
glycolipids role
for immune responses
cell recognition
attachments lipids
importance of membrane fluidity
-lipids diffuse laterally
-proteins not involved in anchoring also diffuse
-proteins need to transmit signals
-transport across by diffusion or via transporter
-vesicles need to bud off and fuse
measuring rate of lateral diffusion in membrane
-membrane with flurophobes
-intense light bleaches flurophobes
-rate of diffusion of flurophobes can be measured
how fluid is the membrane
-biological membranes have constant movement within the bilayer
-rate of membrane lipid movement can be measured
lateral diffusion per second
2 micrometre Per second (a length of the mammalian cell)
transverse diffusion flip flop once every 3 days (rare)
proteins are similar but generally move slower
membrane fluidity temperature
too fluid = membrane disordered too much permeability
too solid = gel slows down movement too much
lipid molecules move faster as temperature increases so membrane becomes more
permeable
increases fluidity
-unsatured lipids gives kinks
-short chains allow fewer interactions between lipids
-high temperature
if you cannot intrinsically reg your temp then you must adapt to surroundings
-organisms regulate their lipid composition
-short unsaturated fatty acids predominate at low temp
-long saturated fatty acids predominate at high temps
plants and their response to heat
-plants have sensors in the plasma membrane that detect changes in fluidity
-fluidity increases indicates temp is increasing
-allows the plant to prepare for heat stress
no cholesterol in
plants and bacteria
ethanol increases
membrane fludity
lipid bilayers are
asymmetric
-teh two layers have different lipid composition
-transverse diffusion (flip-flop) once every 3 days (rare)
-proteins known as phospholipid translators (flippases) catalyse the flip-flop event to maintain phospholipids in the correct monolayer
types of integration membrane proteins
-single span hydrophobic alpha helix either C or N terminal can be intracellular
-multi-spanning containing alpha helices , 7 transmembrane helix protein a big family but can have ore or fewer helices
-beta barrel protein forming a pore
membrane topology =
= arrangement relative to membrane this does not change
maintained by hydrophobic and electrostatic interactions
positively charged amino acids interact with
negatively charged lipid head groups
integral membrane proteins structure
-distinguish between transmembrane cytoplasmic and extra-cellular parts
-loops can form binding sites there may be entire protein domains with extra or intra cellularly
ICAM has several extracellular domains
ICAM is involved in cell adhesion
-expressed in cells of the immune system and endothelial cells
-up-regulated during inflammation
-it has 5 extracellular immunoglobulin domains
ICAM has
-5 extracellular immunglobin domains
-single transmembrane spanning helix
-short cytoplasmic tail
porins
-forms a barrel shaped structure with a pore in the centre
-8 beta strands
peripheral membrane proteins
do not interact with the hydrophobic core of the membrane
-can be cytoplasmic or ectoplasmic
-interact with lipid head groups and integral membrane proteins
-interactions are non-covalent
-electrostatic iteraction, H bonds and van Der Waals bonds
palmitylation
one of many types of lipid anchors
peripheral membrane proteins
proteins coanchored to the membrane through hydrocarbon groups
the protein is covalently attached to a hydrocarbon group
they hydrophobic hydrocarbon group inserts into the lipid bilayer
ankyrin and spectrin
-spectrin cytoskeleton protein creating a scaffold on the intra-cellular side of membrane
-ankyrin binds to several integral membrane proteins AND to spectrin
-maintain plasma membrane integrity via the spectrin-actin based cytoskeletal structure
cells are covered in carbs
-only found in ectoplasmic side of membranes
-attached to both lipids (glycolipids) and proteins (glycoproteins)
-the glycocalyx is a network of glycoproteins with mucus like consistency
carbs on cells role
-physical barrier (protects against viruses and bacteria)
-mechanosensing
-possible roles in cell shape
most protein have at least one carb unit few (___) lipids have carb units
exist as either oligosaccharide chains or single sugar residues
10%
glycoproteins usually have
oligiosaccharide chains
lycolipids usually have
single sugar residues
membrane carbs function
-cell recognition communication and adhesion
-this is especially important in immune responses
-distinguishing self and non-self, infection and transplantation
glycans
are on the outside of membranes and attached to either proteins or lipids
what can cross lipid bilayers
-small hydrophobic molecules
-small uncharged polar molecules
-water
-large uncharged polar molecules
-charged ions
-charged polar molecules
transport by simple diffusion
-solute must be hydrophobic to dissolve in membrane
-rate of transport depend on size and hydrophobicity
-rate and direction depend on concentration gradient (transport continues to a dynamic equilibrium)
channels
gated (voltage, ligand mechanical)
carriers
-permeases, transporters or carriers
three classes of active transporters
-P type pumps, phosphorylate themselves during transportation cycle (ion gradients Na, K and Ca)
-F-type pumps - work in reverse using proton gradients to synthesise ATP
-ABC transporters - pump small molecules as opposed to ions
1 active transporters
conformational change in membrane
P-type pump
lysosome need low PH for hydrolytic enzymes to be activated
transport of several ions makes stomach acid
1)CO2 diffuse in from blood
2)Combines with water to form carbonic acid catalysed by carbonic anhydrase
3)bicarbonate exchanged for CL-
4)H+ transported by P pump
5)CL- enters the lumen via a CL- channel
HeLa human cervical cancer cellist tissue culture Henrietta lacks died 1951 aged 31
-it is easy to spot daughter cell pairs it is uncommon to see to see a dividing or a dying cell even in relatively rapidly growing tissues (HeLa cells divide once every 24 hrs)
1665 Hooke published micrographia and described living tissues being composed of
these pores or cells
he later confirmed the observations of Antoine van Leeuwenhoek that there were single cell animalcules
1875 Mayzel
cell division was described carefully by the polish histoligist
-he showed that salamander embryo cells took up aniline dyes that stained condensed structures in the nucleus (chromosomes or coloured bodies)
1870s phases of mitosis and cytokinesis in a mammalian cell
-the still images of the early micrographers could be put into a sequence of phases that described the life history of cell
interphase (G2) FISH the technique
-cells are grown on glass slides, fixed and permeabilised with detergent
-incubated with fluorescent oligonucleotide probes specific for individual chromosomes
interphase (G2) primers
-they hybridise with their targets the the chromosomes become painted
,
-interphase chromosomes occupy their own territories. They dispersed structure allows access of transcription factors to the DNA: the cells are actively making RNA and proteins
micro tubular spindle fibres grow from the region adjacent to the centrosomes : some extend pole to pole and
others attach to chromatids at
kinetochores
1983 Tim hunt
identified the first controller
process about methionine added at t = 0 **
cyclin is a controller
concentrations rise and fall
cell cycle progression is controlled by heterodimeric complexes each comprising
regulatory cyclin that selects the targets and catalytic cyclin dependent kinase that phosphorylates a target protein
multiple cyclins and multiple CDKs
the important principle is that there are G1 complexes which act as G1 controllers
cyclin CDK complexes phosphorylate their targets which prepare the cell for S phase and promote the expression pf S phase cyclin
S phase cyclin CDK complex phosphorylates
its target which control chromosomes replication
The S phase cyclin CDK
complex phosphorylates its target which control chromosomes replication
how is spindle formation activated
G2/M cyclin complexes phosphorylate their targets
one CDk adapted by different cyclins led to the idea
if this was an evolutionary intermediate and investigated by NURSE in 1996
marine alga O. tauri a photosynthetic machine had
genome sequenced 2004-2006 and there are two cell cylcle cyclins only
the G1 D and G2B
fission yeast Sz
1996 Genet a quantitive model for the ccdc control of S phase and mitosis in fission yeast
2010 found answer that the driving of the cell cycle with a minimal CDK control network
looking at Sz pome survive with one CDK
1)the CD13 and CDC2 genes were fused with an in frame linker
2)expressed in Sz. Pombe
3)all other cyclins and CDKs were deleted one by one checking for viability each time
implications:a primoridial cyclin and a primordial CDK were sufficient to control the cell cycle and gene duplication of cyclin or of the CDK allowed new combinations to perform at different stages ion the cell cycle;e
resulting in complexity seen in modern oranisms
cells can leave the cell cycle and enter a
quiescent phase
What can a cell do?
-not too much room for division uncontrolled proliferation = cancer
If these G0 arrested cells then re-enter the cell cycle a
tumour may reform
Activation of growth factor receptors
they start at the plasma membrane by binding of specific ligands (EGF epidermal growth factor)
Activated EGFR is bound by adaptor molecules that
recruit and activate the cytosolic membrane bound ras
Biological significance the signal has been transducer from the extracellular side of the plasma membrane to the intracellular side of the membrane
Signal transduction via an enzyme cascade
Amplifies the signal
growth factor summary
highly conserved signal transduction pathway. Stimulation of growth factor receptors results in Raw recruitment, signal transduction from Raf to MAPK and expression of G1 cyclins and CDKs that return to G0 cell to G1
what can go wrong
causing unregulated prolifearation
tumour suppressor genes arrest the cell cycle in
G1 some in G2
they suppress G1 by
slowing down entry into S phase
giving time for DNA to repair following mitosis
key regulator is pRb(retinoblastoma protein)
how does pRb work?
S phase proteins are under the control of E2F transcription factors in G1 pRb binds E2Fs and inactivates them so the cell cannot make S phase proteins until
the G1 cyclin /CDKs have accumulated enough to phosphorylate Rb and inactivate it allowing S phase proteins to be made
p53 another tumour suppressor protein
loss of protein creates genome instability resulting in aneuploidy
TP53
tumour suppression is severely reduced if damaged
associated with 50% of human cancers
DNA damage, hypoxia and other stresses
cell cycle arrest, DNA repair and cell cycle restart
or apoptosis death of damaged cells
cellular and genetic stability
human papilloma virus inhibits
p53 activity and others* check lecture 27 for specifics
necrosis
cells that die through tissue damage exhibit different morphological changes
dying cells swell and burst and intracellular contents are released into extracellular milieu
causing inflammation
2002 Nobel prize Brenner, Horvitz and Sulston
for their discoveries of genetic regulation of organs development and programmed cell death
apoptosis
dying cells shrink
capase
cleaves target at just the C terminal leading to cell death
normally kept inactive by trophic signals from neighbouroring cells
therefore apoptosis is our natural state
internal triggers for apoptosis
recognition of irreparable DNA damage by the p53 pathway
DNA damage from radiation, chemo mistakes in replication and lack of trophic signalling
external triggers for apoptosis
-recognition of stress
-heat, radiation and starvation
can damage cell membranes leading to apoptosis
developmental triggers for apoptosis
apoptotic programs remove the webbing between our fingers during foetal development and remove a number of neurons
GFP tagged proteins
help see localised sub cellular structures
how do proteins reach their destination
sorting signals
they are part of the protein
-short peptides at N or C terminal = can be removed after use or kept on if needed again
-3 dimensional domains (secondary tertiary structure) = for transport like lysososomes
-other molecules attached to the protein like post-translational modifications of sugars and lipids
what happens to sorting signals
-they are recognised by specific receptors
-which in turn trigger transfer of client protein to correct destination
-every organelle uses different receptors and different sorting processes
-if any of this processes goes wrong the cell is in trouble
models of protein transport-gated transport
import into and export out of the nucleus
models of protein transport-transmembrane transport
protein import into ER and mitochondria uses transient translocation channels in the correct membrane
models of protein transport-vesicular transport
secretion along the organelles of the secretory pathway we will cover targeting to lysosomes
gated transport into the nucleus
proteins and other macromolecules move between the cytoplasm and nucleus via large aqueous nuclear pore complexes
a mammalian nuclear envelope contains 3000-4000 nuclear pore complexes
-side view and face-on EMs of nuclear envelope showing the distinct 8 fold symmetry of nuclear pores
small molecules (5kDa or less) can
rapidly diffuse between cytoplasm and nucleoplasm
proteins of 20-40,000 Da
diffuse more slowly
proteins >40kDa cannot enter
and RNA/ribosomes cannot exit unless they carry nuclear localisation or export signals
22% of human proteins are
nuclear and must be imported from cytosol
diffusion barrier is caused by
unstructured regions of NPC proteins forming tangled network blocking the passive diffusion of large molecules
NPC is
125 x 10^6and has 30 different nucleoporins`
nuclear localisation signals (NLS)
rich in lysine and proline and can be in any position of the passenger (cargo) proteins
so long as they are exposed to the surface of the protein
importins are the NLS receptors
recognised by a family of cytosolic nuclear import receptors each member being responsibsible for a set of cargo molecules
nuclear import
since transport is through large pores, fully folded proteins and newly formed ribosomal subunits can be transported in and out respectively
the importing binds the NLS and the FG repeats in the FG nucleoporins of the fibrils and channel
how does importing know when to let go of its cargo
-GTPase switch
-a protein that can either bind GTP or GDP and can hydrolyse GTP to GDP
and assumes a different conformation depending on which nucleotide it binds
-different conformation = different activity
Ran-GDP has conformation
A and in this state only in cytosol
Ran-GTP
has conformation B and in this state is only in nucleus
Ran-GTP arriving in cytosol is
immeditly converted
extracellular matrix (ECM)
supports cells and their influences
-strength
-elastcity
-turgor
Gl;ycosaminoglycan (GAG)
chains are repeated units of negatively charged dissacharides that form linear chains
they attract cations(Na+) causing large amounts of water to be sucked into the matrix
they occupy a large volume relative too their mass and creat turgor
negatively charged attracts cat ions so
osmosis occurs
an aggrecan aggregate
Is composed of aggrecan bound to hyaluronan
-a proteoglycan with 100+ GAG chains
aggrecan molecular weight
2.10^8
proteoglycans(PGs) that contain GAGs
means pore sizes vary
they can bind signalling molecules or can bind to proteases to concentrate them
fibroblasts in the ECM
what you see mostly is fibrillar collagen
about 25% of protein in body is collagen
fibrillar collagen structure
N terminal polypeptide (150 aa)
signal peptide(SP) directs the nascent protein
glycine residues (the repetitive structure promotes trimerisation)
-C terminal propeptide(250 aa)
modifications in ER
1)about 50% of the proline residues are hydroxylated (-OH) by collagen propel-hydroxylases this requires vitamin C
2)specific lysine residues are hydroxylated this also requires vitamin C
3)the c terminal domains are N glycosylated
4)and mostly in the Golgi some of the hydroxylsysl are O glycosylated by addition of galactose or glucose galactose
winding production triple stranded helical structure
with highly soluble N and C termini
function of N and C termini
-one other function is to keep trimers apart to stop them forming fibrils
-until late the secretory pathway when the pro peptides are removed forming tropocollagen
fibripositors
individual tropocollagen are cross linked to form fibrils
the type IX collagen is a fibril-associated collagen that can interact with type II
this interacts with type 1 collagen fibrez
elastin
a hydrophobic elastic protein with extensive crosslinks
highly prevelant in ECM of arteries gives tissue their elastcicty
cytoskeleton
-provides mechanical strength
-drives organelle movement
-severs as an anchor for cell-cell junctions
-drives chromosomes segregation in mitosis and splits the cell in two(cytokinesis)
-enables cell movement and muscle contraction
microtubules
non covalent heterodimer of alpha and beta subunits
-both subunits bind GTP
-there is polarity a += end and - end
-GTP bound heterodimers bind only at ends growth of microtubule is unidirectional from the + end only
-microtubules are stiff stuctures
growth of micro
-grows/shrinks only at + end
-they are dynamically unstable
actin filaments
-non covalent polymers of actin monomers
-each monomer binds ATP (which is replaced by Ap in the filament)
-two filaments twist around each other to form an actin molecule
-there is a polarity a plus and minus end
IFs are rope like fibres ,Ade of intermediate filament
proteins that are made of alpha helical monomers that associate into coiled coils that then associate in filaments
-IFs form a very large diverse family of non-nucleotide binding proteins including the nuclear laming and epithelial keratins
-intermediate filaments function = they provide mechanical strength they bend but do not break
types of junctions
**
adhering form
HOMODIMERS WITH THE EXTRACELLULAR PART OF EACH POLYPEPTIDE FOLDED INTO FIVE CADHERIN REPEATS
THERE ARE CA 2+ BINDING SITES BETWEEN EACH BPAIR OF REPEATS
cadherins and homophillic binding
-interact weakly so each pair is disassembled
Wilson HVP (1907)
-on some phenomena of coalescence and regeneration in sponges
-they were disaggregated into single cells by forcing pieces of sponge through a fine sieve
-tried to make a hybrid from two different species by disaggregating cells and mixing them but no hybrids so sponges have HOMOTYPIC recognition
adherent junctions summary
-the cell to cel interacting membrane proteins are cadherins
-the homophilllic interactions they make are weak, but many acting together makes for strong attachment proteins
-enable cells to be recognised
-junctions connect indirectly to actin cytoskeleton and influence the co-ordinated contractions of cells
-complex interplay between chemical signalling pathways and cell-cell adhesion
integrins
transmembrane proteins that allow the internal cytoskeleton to grip onto molecules of ECM when necessary
integrins
transmembrane proteins that allow the internal cytoskeleton to grip onto molecules in ECM when necassary
interns are receptors for ECM molecules
-a alpha/beta heterodimerthat links an ECM to talin
-talin links to actin other proteins reinforce the linkage
-integrins there are 8 different beta chains and 18 different alpha chains each with distinct ligand binding prop[erties `
junctions
**
