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what are the two mechanisms of cellular control

changing the specific activity of enzymes

changing the concentration of enzymes by synthesis and degradation


discuss 3 methods of changing the specific activity of enzymes

a)non covalent boning. allosteric sites

b)covalent bonding.
usually done by phosphorylation; the phosphate group has a negative charge and therefore changes the structure of the enzyme

c)protein-protein interaction)


enzymes are often phosphorylated to change their specific activity, where does phosphorylation occour and where does the phosphate come from

the phosphate comes from ATP, phosphorylaiton id performed on amino acids with a hydroxyl group (serine, tyrosine)


why are yeast used to study the cell cycle

the genome is small, so cell cycle only takes 2 hours (can take up to 24 in other eukaryotes)
their small haploid genomes are easy to clone and select mutants from


saccharomyces cerevisae is the usual model organism yeast, but another yeast species is used to study the cell cycle, which species is it and why is it used

SCHIZOsaccharomyces pombe. saccharomyces cerevisae is a budding yeast and doesnt divide down the middle


what are the CDC genes

the cell division cycle genes which regulate cell division


why is studying cells with mutant CDC genes hard

these genes control the cell cycle, when they are mutated the cell dies. some mutants can be studied (those that are temperature sensitive)


name the 3 'checkpoints' where the cell commits to certain next steps

in G1 the cell commits to DNA synthesis
in G2 the cell commits to mitosis
in M phase the cell commits to complete mitosis


discuss what Cdk's are

cyclin dependant kinases are enzymes that initate many important parts of the cell cycle but they only work when cyclin protein is present to activate them


how does the G1 to S phase transition occour

G1-cyclin accumualtes and activates G1-Cdk, which activates transcription of G1/S cycllin.
G1/S cyclin accumualtes, which activates G1/S-Cdk, which increases activity of S-Cdk, which initiates DNA synthesis

the three ways that G1/S-Cdk increases activity of S-Cdk are;
1)activating transcription of S-cyclin
2)inhibiting the S-Cdk inhibitor
3) inhibiting the D-Cdk ubuiquitin ligase which would normally causes S-Cdk to be degraded


how does entry to mitosis occour

M cyclin is synthesised which activates M-Cdk.
initially, M-Cdk is inactivated by Wee1 kinase, but when DNA replication is complete, polo like kinase activates cdc25 phosphatase, which dephosphorylates and activates M-Cdk
M-Cdk activity causes entry to mitosis (nuclear envelope breakdown and chromosome condensation caused). M-Cdk inactivates wee1 kinase and activates more cdc25 phosphatase (positive ffedback)


discuss proteolyis in the cell cycle

degradation of cyclin has massive impacts on weather or not the cell cycle continues


define protein turnover

the balance betweeen protein synthesis and degradation


discuss proteasomes structure

a large hollow cyclinder shaped protease (enzyme that degrades protein)


what structure of a proteasome stops trandom degradation of unwanted proteins and how

the 'cap' which stops other proteins entering which dont have a ubiquitin protein attached to them


what is the protein added to a protein which makes it destined for degradation? which enzyme adds the protein? where is the protein added to the protein? how does the enzyme know which proteins to add the protein to

ubuiquitin, ubiquitin ligase, lysin AA side chain; ubuiquitin is added to proteins with degradation signals on them (abnormal structure)


name the chemicals that signal for cell division (entry to cell cycle) and mass increase

mitogens stimualate cell division and growth factors stimulate mass growth


how to mitogens stimulate cells to divide

mitogens bind to the cell surface and cause secondary messenger action which causes transcription of G1-cyclin and transcription of enzymes which inhibit/degrade G1-inhibitor proteins


name two examples of mitogens

platelet derived growth factor (PDGF); stimualtes many cells to divide

erythropoietin; stimulates RBC precursor cells to divide


define cancer and discuss retinoblastoma

cancer in unregulated growth and cell division

retinoblastoma cancer occours when the RB1 gene is mutated (normally would be expressed into RB1 protein which prevents premature cell division by stopping transcription of G1/S-cyclin and S cyclin and therefore blocking entry into the S phase)


what is the RB1 protein

it is a gene that prevents premature entry to mitosis by blocking entry into the s stage (inhibts synthesis of G1/S and S synthesis.
RB1 mutation lead to retinoblastoma.


discuss the mechanism by which cells with mutated DNA have replication paused until DNA is repaired

p53 would normally be ubuiquinated and degraded (low concentration exist and replication occours)
when DNA is mutated, phosphorylation of p53 occours and the ubuiquitin protein cant be added; p53 accumualtes, activates transcription of p21 protein which accumulates and inhibits activity of G1/S-Cdk and S-Cdk.


discuss how mutated p53 leads to many human cancers

p53 would normally prevent replication of mutated DNA occouring, if its mutated itself then DNA will be replicated when it shouldnt


state the stages of the cell cycle and which stage takes the longest

G1; protein synthesis
S; DNA synthesis (12 hours)
G2; checking DNA
G0; resting state


describe how sister chromatids are seperated during anaphase

the seperase protease enzyme breaks down the central cohesin protein complex holding the sister chromatids together

M-Cdk activates a ubuiquitin ligase (APC) which ubuiquitinates securin which is then degraded (the enzyme that would normally inhibit sperase)


why does exit from mitosis occour

APC (a ubuiquitin ligase) ubuiquitinates M-cyclin, which is then degraded.
lack of M-cyclin inactivates APC


define necrosis

rapid accidental cell death, usually for cells which are injured


discuss the processes involved in apoptosis

cell shrinkage
nuclear membrane blebbing
cytoskeleton breakdown
phagocytosis by phagocytes


what causes phagocytosis of cells during apoptosis

alteration of the plasma membrane; proteins placed on membrane


what is the principle protease used for apoptosis

what element does it have at its activate site

where does it cleave

what is the name of the inactive precursor that is synthesised before the enzyme is needed

how does this precursor enzyme become activated


sulphur at active site

cleaves at aspartic acid amino acids

procaspases are the precursor fort he caspase

1) inhibitor caspases cleave procaspaces between their large and small sub unit as well as cleaving off the prodomain.

2)the large and small subunits rearrange next to eachother

3)dimer formation between two complexes

4) these are known as 'executionar caspases' cleave more procaspases and break down proteins


how are DNases activated

an executioner caspase breaks down the DNase inhibitor protein


how is apoptosis initiated via external signals

1)foreign pieces of protein are bound to the MHC (major histocompatibility complex) on the plasma membrane

2)a cytotoxic T lymphocyte binds to MHC by its T cell receptor (TCR), recognises the foreign compound and activates apoptosis

3)the target cells Fas receptor binds to the T cells Fas ligand protein on its surface

4)all fas receptors on the surface aggregate and come together, and so do the associated initator procaspases (8 and10) due to the binding of the FADD adaptor protein

5)when procaspases are brought together they activate eachother which the activate executionar caspases


why does Fas aggregation cause procaspases to come together

Fas receptor has a death domain which FADD adaptor protein binds to, and FADD adaptor protein has a death effector domain which prcaspases 8 and 10 bind to


what is the name of the complex when Fas receptors aggeregate and bring together the procaspases and FADD adaptor proteins

Death inducing signal complex (DISC)


discuss internal initiation of apoptosis

1) mitochondria releases cytochrome C
2) cytochrome C binds to adaptor proteins called Apaf1
3)apaf 1 group together into an apoptosome via CARD domains
4)procaspase 9 come together and bind via their CARD domain and activate eachother
5)procaspase9 activates executionary procaspases

mitochondria release cytochrome C due to an inhibitor inhibiting Bcl2 genes, bax and bak proteins form pores in the membrane


discuss how cytotoxic T cells cause cause internal initation of apoptosis

1)T cell releases perforin which creates holes on plasma membrane
2)granzyme B enters cell and cleaves bid protein
3) tBid enzyme activates bax and bak
3b)granzyme also acitvates procaspase3 and 7


how is apoptosis prevented

survival factors like insulin like growth factor 1 (IGF1) inactivate BH3 which would normally activate bax and bak pore formation by inhibiting their inhibitor

IGF1 activates anti-apoptotic genes; no cytochrome C release


what is p53 and how it relate to apoptosis

a protein that activates both internal and external initaitons of apoptosis; it activates BH3 pro apoptotic proteins and activates transcription of Fas proteins


when does protein folding occur

whats the name of the intermediates that form during folding

give an example of incorrectly folded protein

as the protein is being synthesised

a protein with hydrophobic acids on the outside of the portein is a mutation; proteins are sticky and aggregate with other proteins

proteins form molten globule intermediates during synthesis


what is the function of chaperone proteins

what are the two families of chaperones

chaperones bind to incorrectly folded proteins, stop them from aggregating and catalyse refolding, or send proteins to proteases for degradation if damage is irrepairable


discuss the two families of chaperones

what does hsp stand for

when do these proteins increase in concentraion

hsp70 are those that bind to exposed hydrophobic regions DURING synthesis

hsp60 (part of the chaperonin group) are those which have a cavity and provide a space for protein refolding AFTER synthesis

heat shock protein

increase in concentration when cells are exposed to high temperatures


discuss the E coli and eukaryotic cytosolic hsp60

state a similarity between the two chaperones

e coli; GRoEL with a removable GroES cap

eukaryotic; TRiC/CCT, which has an inbuilt lid which is good because part of multidomain proteins can remain outside the cavity

both have 2 cavities but only one can be used at once


describe the method used to move proteins

a) from the nucleus to the cyosol

b) from the cytosol to organelles

c) from organelle to organelle or from organelle to the plasmam membrane

proteins move from the nucleus to the cytosol via nuclear pores (gated transport)

proteins move from the cytosol to organelles via transmembrane transport

proteins move from organelle to organelle or to the plasma membrane via vesicular transport


what is the name of the polypeptide sequence that specifies the proteins destination

signal sequence,

can be at amino terminal, carboxy terminal or middle of protein. Amino terminal is common

is sometimes removed before it reaches its destination


discuss the fucntion and location of the endoplasmic reticulum

a continuous network with the nucleus used for protein synthesis


discuss the name and characteristic of the signal sequence that directs the polypepeptide to the endoplasmic reticulum

signal peptide, which is a 20 amino acid long sequence, usually present at the amino terminus.
the signal peptide has no clear consensus sequence but has a distinctive feature which is 10 hydrophobic amino acids in the middle of the sequence


discuss how proteins are synthesised and directed towords the ER

1) in the cytosol, the mRNA begins being translated by ribosomes, and when the signal peptide is tranlated, it binds to a signal recognition particle (SRP) which moves the whole complex to the cytoplasmic reticulum where the SRP binds to the SRP receptor

2)The signal peptide is inserted through the translocator pore and the polypeptide grows inside the ER lumen

3)the signal peptide is cleaved, releasing the protein


describe what happens to proteins that enter the ER lumen

they have a oligomer (14 sugars) added to their asparagine amino acid by the enzyme Oligosaccharyl transferase, which occours as the protein is being synthesised


what is the oligosaccaride attached to as its being synthesised

a transmembrane protein in the plasma membrane called dolichol phosphate


discuss the way the oligosaccaride is attached to the polypeptide

it is N linked; attached by the N atom of the sugar molecule


apart from being glycosylated, what is something else done to proteins in the ER lumen

protetion folding by an Hsp70 chaperone called BiP

calnexin and calreticuin assist with mis folded proteins and recognise them because they have one glucose added to their oligosaccaride


what is removed from the proteins oligosaccaride which signals for protein export and degradation

removal of the 2 terminal mannose molecules


discuss the structure of golgi apparatus

a series of sacs called cisterna, each of which has a specific function.
the sacs the protein goes through first are termed 'cis' sacs and those travelled through last are termed 'trans' sacs


discuss three ways the protein is modified in the golgi apparatus

1) phosphorylation; occours to those destined for the lysosome

2)removal/addition of sugars

3) sulfation of amino acids and carbohydrates


what are the theories used to explain how golgi apparatus operate

cisterna are fixed and protein move from sac to sac in vesicles

cisterna move and change activity while backwards transport of vesicles occours to balance the forward movement of materials


what is the name of the process by which vesicles bud off and fuse with membrane

exocytosis endocytosis


what is the name of the structure that sororunds most vesicles

protein coat


discuss the COP1, COP2 and clatherin coats that sorround the vesicles

COP2 coated vesicles are used to transport proteins from the ER to the GA

clatherin is used to move proteins from GA to lysosomes

COP1 is used to move proteins from the GA to the ER


discuss the interaction between proteins on vesicle membranes and those on target cell plasma membranes which pull membranes together

the V-snare protein vesicles binds tightly with the T-snare protein on the target cell

the Rab protein (GTPase) on vesicles interacts with Rab effectors on target cell

those are known as tethering proteins


what is the point of glycolating proteins

these can be used for cell to cell interactions like recognitionand adhesion

protection; extracellular matrix

production of mucus (micins are glycoproteins)


what is the name of the proteins that should reamain in the ER but ma leak out due to poorly understood signals

how do they leak out

Cargo proteins

may bind to proteins on the vesicle protein coat


name the sequences proteins have if theyre sopposed to remain in the ER

KDXX (membrane protein)

kDEL (soluble protein


how are cargo proteins returned

they bind to receptors like the KDEL receptor and are then transported back to the ER in COP1 vesicles


what amino acid does K represent



discuss how rab efffectors use GTP

inactive rab-GDP in cytosol binds to the source membrane, and exchanges GDP for GTP, moves to target membrane, binds to the rab effector and leaves the membrane once GTP has been hydolysed


proteins that should remain in the endoplasmic reticulum have specific sequences (KDEL, KDXX) where are thse sequences located

carboxy terminus


what are the two types of protein that should remain in the ER, what carboxy sequence do they have

membrane protein; kDXX

soluble protein; KDEL


what are the two types of glycsolation that occour in the golgi apparatus

N linked; builds of glycsolation already taken place in ER, on the asparagine AA

O linked;sugar molecules added to the serine and threonine AA's


what are three pathways by which proteins are moved from the golgi apparatus

1)movement to the lysosomes. acid hydrolyases have a phosphate added to their mannose, the protein then binds to M6P receptor and moves to lysosome in a clatherin coated vesicle

2)constitutive secretion pathway; movement of vesicles tplasma membrane

3) regulated secretion pathway, vesicles dock with plasma membrane but arent releeased intil an extracellular signal causes them to be. food is eaten which signals for the pancreas to releas degradative enzymes into the gut


discuss variety in mitochondria genomes

the genome is varied between kingdoms but within a kingdom species have similarly organised mitochondria


how many copies of mitochondrial genome are in each cell

many; there are many mitochondria per cell


how is mitochondrial DNA inherited



what are some abnormal features of mitochondrial DNA

extensive RNA editing
abnormal genetic code


how many gene coding proteins do eukaryotes have, what do these genes code for

5 to 60
they code for membrane enzymes that are needed for electron transport and ATP synthesis, ribosomal RNA


recall the michaelis mention equation

V=kmax x s/Km + s


discuss the differences between first order and second order reactions

first order reactions have one reactant, second order reactions have 2 reactants


what are the assumptions of michaelis menton kinetics

only occours if theres an excess of substrate

if the concentration of P is low then the back reaction is insigignificant

only applies if one substrate concentraion is varied at a time


discuss the difference between reversible and irriversable enzyme inhibitors

reversable inhibitors bind and then dissociate
a)competitive inhibitors compete for the active site, dont change Vmax but increase the Km

b)non competitive inhibitors bind to sites other than the active site, decrease the Km but the Vmax remains unchanged

c)mixed inhibitors effect both the Km and Vmax

irriversible inhibitors bind covalently and dont dissociate


what must a competitive inhibitor have to be to be competitive

present at a higher concentration than the Ki (how tightly the inhibitor binds to the substrate)


what is the intermediate molecule formed during an enzyme reaction

what does formation of a transition state require

how does an enzyme speed up rate of reaction

transition state

requires activation energy]]decreases activation energy required which makes rate of reaction quicker


how does the enzyme lower activation energy

orienting substrates in a way that encourages a reaction to occour between them

rearrangeing the electrons in the substarte to create positive and negative charges that encourage a reactin to occour

straining the substrate and forcing it into a shape closer to the transition state

forming covalent bonds with the substarey


discuss some charcateristics of the active site

the substrate fits tightly into it

often span multiple domains (polypeptides)


what molecule is often bound to the active site cleft and helps with catalysis



what is one example of a coenzyme

zinc ions


what is the function of lysozyme

it is a defensive enzyme which performs hydrolysis of the petidoglycan compound found in bacterial cell walls


discuss how lysozyme lowers activation energy when hydrolysing peptidoglycan

it strains peptidoglycan and changes its conformation


which amino acid in lysozyme donates a proton to sugar bonds in peptidoglycan and which amino acid then breaks the bond between 2 sugars

glutamtic acid35
aspartic acid 52


what happens if there is a mutation to the glutamic acid35 and aspartic acid52

the enzymes activity is greatly reduced


what is the rossmann fold

a binding domain for NAD+ that is present in many dehydrogenase enzymes


where does cyclin bind to Cdk enzymes

binds between an alpha and a beta domain, pushes them apart and exposes an ATP binding site


apart from synthesis and degradation of cyclin, what also changes Cdk activity during the cell cycle

phosphorylation and dephosphorylation of Cdk


which genes prevents premature cell division

RB1 gene/protein. stops stranscription of G1/S cyclin and s cyclin so cell doesnt enter DNA synthesis stage


what is the name of the primary hsp70 in the ER lumen

which two chaperons help it with misfolded proteins,

how do these 2 recognise mis folded protein


calnexin and calreticuin

glucose added to end of the oligosaccaride