Biochemistry Flashcards
What is miRNA?
binds to mRNA - promotes degradation/inhibiting translation
what is snRNA
Involved in RNA splicing
What is the function of holoenzymes?
Act as a primer for RNA polymerase
What is a Pritbrow box, and what does it act as?
is a promoter region comprised of high A-T concentration
What does a termination sequence look like in terms of bases?
G-C rich region followed by an A-T rich region
What is the mass of each prokaryotic ribosomal subunits and the total ribosome mass
50s and 30s
total is 70s
At what codon does translation initiation commonly occur?
AUG
What 2 amino acids cannot be included in an alpha-helix structure and why?
Glycine - too flexible for the structure
Proline - too kinky because its angle it 10 degrees off for the optimum structure for an alpha helix structure
What is the optimum angles for Psi/Psh bonds to be for an alpha helix?
60
What is the purpose of randomly coiled parts of a polypeptide chain?
allows ligand binding and acts to connect different different protein domains
Function of beta-ME?
breaks di-sulphide bridges in tertiary-structures
How did Anfinsen fully unfold a protien?
8micromililitres of urate
excess beta-ME
What are the 4 driving forces of protein folding?
hydrophobic affect
Pi-Pi bonds between aromatic rings
electrostatic interactions
hydrogen bonds
What amino acids are phosphorylised in a quaternary structure?
serine/threonine/tyrosine
What is acetylisation and how does it do it?
compensates for the positive charge of lysine by forming acetyl-lysine
What is palmitoylation and what amino acids are involved?
long lipid groups help anchor proteins in the membrane
glycine, cysteine, lysine
How does AAT deactivate elastase?
AAT lures in elastase then, rotates it 180 degrees
What is the function of scaffolding proteins?
allows other proteins to bind to for compartmentalisation
also used for cell signalling
What is a glycosaminoglycan
amino acid within a polysaccharide
Proteoglycan function?
holds together protein placement in connective tissue
What are peptidoclycans formed from?
polysaccharide-peptide complex
alternating N-acetylglucosamine and N-acetylmuramic acid
In bacterial cell walls What forms the bridges between the amino acids in the structure?
penta-glycine bridges
What is the defintion of a ligand?
a molecule that binds to a macromolecule to form a complex which initiates a conformational change to generate a signal
What is a juxtacrine signal?
A signal that is only between 2 neighbouring cells
What is an autocrine signal, and what is the distinction between intracrine signal?
an autocrine signal is one where the signalling cell and the target are the same cell
intracrine signals are a type of autocrine signal but the signal remains within the cell
What does NO cause?
chronic relaxation
What type of signal is hydrophobic but cannot diffuse into a cell, how is it transported into the cell and where is its receptor located?
thyroxine signals
transporter proteins
in the nucleus
What are ligand-gated ion channels composed of?
4 polypeptides, 3 hydrophobic, and 1 aliphatic, lines the pore
What happens when G-protein releases GDP and gains GTP?
alpha-G dissociates from beta-G and the both become active
What are the functions of activated G-aplha?
Activates adenyl-cyclase
can also deactivate adenyl-cyclase
What does calcium bind to for intracellular messaging, and what does that bind to?
calmodium, then CaM-kinase
what is the function of calmodium-dependent kinase
regulates many proteins activity
How does protease modify an enzyme?
cleaves a peptide chain on the active site
how does a kinase modify an enzyme and how is this reversed?
the kinase can phosphyrale the enzyme
reversed by any phosphotase
what is a K-type modification?
an enzyme binds to a substrate to increase the affinity of that substrate for another enzyme
define a plasmid?
small circular DNA found in bacteria
what DNA polymerase can be used for PCR
Taq-DNA polymerase
outline the function of restriction enzymes?
detect the specific sequence of DNA to cut
what are the required components for PCR
ds DNA template
pair of oligonucleotide primers
DNA polymerase
Nucleotides
Why don’t restriction enzymes cleave bacterial DNA
ECO RI methylase methylates the DNA sequence which prevents restriction enzymes from cleaving the sequence
why does ECO RI methylase only methylate bacterial DNA?
when already methylated DNA undergoes semi-conservative replication, there is a strand that is not methylated and one is
ECO RI only methylates hemi-methylated DNA, so it methylates the other strand and wont methylate non-bacterial DNA
What is the difference between type I/II/III restriction enzymes?
Type 1 cuts DNA 1000 base pairs away from its recognition site
Type II cuts DNA at the recognition site
Type III cuts DNA 25 base pairs away from the recognition site
Function of DNA ligase
catalyses phosphodiester bonds and used to stick DNA to a plasmid
what do you do when the gene and the vector do not contain compatible enzymes
use nuclease to create blunt ends that can be inserted into the plasmid
use DNA pol to fill in ends of the gene to create a blunt end
how do you prepare a competent cell for a plasmid, and what is the purpose of the second step?
50mM CaCl2
heat shock of 42C for 2 minutes to increase membrane fluidity so the plasmid can pass through
how do you calculate specific enzyme activity and what does it indicate?
enzyme activity/total amount of protein
indication of purity
What is Vmax and how do you find out the Km
V-max - maximum possible rate
Km - half of Vmax on a V-[S] graph and do a line from half the Vmax to the line, and then go down to the x-axis, and thats the Km
what is Kd, and what does a small Kd indicate?
Kd is the affinity for substrate and the lower the Kd the higher the substrate affinity
outline the micheales-menten model
E + S ES E + P
K1/K-1 Kcat
how does a competetive inhibitor affect the Km and Vmax?
Vmax stays the same, Km is larger
how does a non-competetive inhibitor affect the Km and Vmax?
Vmax is smaller, Km stays the same
how do uncompetetive inhibitors affect the Km and Vmax
both Vmax and Km are smaller
what are some enzyme strategies to reduce the activation energy and why do they work
destabilise of substrate and stabilisation of the transition state
basically makes it more energetically favourable for the substrate to enter the transition state
what amino acids form a catalytic triad
serine, aspartate, histidine
what are is the main cause for Turner/Klinefelter syndrome
non-disjunction - chromosomes fail to seperate for gamete formation
what is philadelphia chromosome
translocation of material between chromosome 9 and chromosome 22
what is cystic fibrosis
when chloride ions don’t move outside the cell, causing mucus to build on top of it
what causes sickle cell disease
A is changed to T at position 17 for the beta-chain of haemoglobin.
changes glutamic acid to valine
key features of B-DNA
right-handed
10 base pairs per turn
3.4nm per turn
2nm in diameter
features of A-DNA
right handed
formed when DNA is dehydrated
wider, tilted, offset from its axis
features of Z-DNA
left handed
formed by high G-C concentration
at what nanometre does DNA absorb maximally
260nm
difference between sucrose density centrifugation and cesium chloride density gradient centrifugation
sucrose density - not an equilibrium method - everything eventually sinks
cesium chloride - is an equillibrium method - material floats in its corresponding position of density
what makes the primer and what is it made from
primase
made of RNA
what are the short fragments of DNA from the lagging strand called and what joins them together
okazaki fragments
DNA ligase
what does the missing OH mean for dideoxynucleotide triphosphate mean in terms of PCR
can be added to the DNA chain but causes chain termination because a nucleotide (e.g. dATP) cannot bind to it
what dye is used to visualise DNA strands in agarose gel
ethidium
what is added to polyacrylamide and why, what is used to visualise the DNA fragments
8M of urea to seperate single stranded DNA
autoradiography/addition of a fluorescent group via covalent bonding
what is the difference between the sanger and next generation methods in sequencing
in the next generation sequencing method an image is taken after addition of each nucleotide
reversible terminator cleaves off the fluorescent dye allowing addition of another dNTP - continuing PCR
what is the difference between DNA and RNA
RNA contains ribose sugar
contains an extra OH group at the 2’ carbon
what are promoters and where pribnow boxes located
sequences that determine the start for transcription
prokaryotes
how fast does transcription occur
50 bases per second
how many bases does RNA polymerase bind to
30 bases
how is RNA modified to form tRNA
some bases and sugars are modified
CCA is added at the 3’ end
difference between RNA I/II/III in eukaryotes
1 - involved in rRNA
2 - involved in mRNA
3 - involved in tRNA and 5S rRNA
why is a poly-A tail added to the 3’ end of pre-mRNA
increases stability and specifies it for export into the cytoplasm
what is the ‘cap’ added to pre-mRNA
a 5’-5’ triphosphate link to 7-methyl-G
function of fibronectin mRNA and what synthesises it
is an mRNA that include EIIB and EIIA
produced by fibroblasts
what does abnormal processing of beta-globin in RNA transcript cause
anemia due to defect in haemoglobin synthesis
what is the difference between constitutive and regulative expression
constitutive - genes expressed all the time
regulative - genes expressed under certain conditions
difference between inducible and repressible gene expression
inducible - molecule binds to repressor protein to remove it from the operator region to activate transcription (lactose)
repressible - bind to repressor protein so that it binds to the operator region (tryptophan) aka a corepressor
difference between promoter and promoter-proximal regions
promoter - common in most genes (TATA box)
promoter-proximal - specific for particular gene
difference between basal and regulatory transcription factors
basal - bind to promoter regions
regulatory - bind to proximal-promoter regions
function of streptomycin
freezes initiation complex and causes misreading in mRNA
function of tetracycline
prevents binding of incoming amino-acyl tRNA
function of erythromycin
binds to 50s sub-unit and prevents translocation
function of puromycin and why can’t it be used as an antibiotic
mimics the terminus of amino-acyl tRNA so is added to onto the chain-causing premature termination
not selective to ribosomal ribosomes, will fuck up eukaryotic ribosomes
why can there be no rotation around the planar peptide bond
resonance gives a partial double bond
definition of protein domain
area of a protein that folds independently
what enzymes acetylate and deacetylate lysine
acetylate - KAT’s
deacetylate - KDAC’s
what is a zymogen
precursors of enzymes that are activated after a proteolytic cascade
when is PKR activated and by what and what causes this and what is that thing a sign
activated when double stranded RNA is detected - a sign of infection
activated by dsRNA
function of porphryin rings
prosthetic group in proteins involved in coordinating transition metals
what is Fe2+ coordinated by in haemoglobin
histidine residues
effect of CO2 on haemoglobin
CO2 dissolves in water to form carbonic acid
decreased pH leads to protonation of histidine in beta-haemoglobin
causes the chain to be more likely to enter T-state
Bohr effect
structure of hemicellulose
beta 1-4 xylopyranose
structure of pectin
mixture of branched polysaccharides
rich in alpha 1-4 galacturonic acid
structure of agaraose
D-galactose linked with beta 1-4 galactose
has an ether bridge
structure of chitin
polymer of N-acetylglucoseamine
function of a GPI anchor
helps binding of proteins to lipids at the lumenal or surface membranes
what diseases are caused by dis-functioning Ca2+/K+ channels
Ca2+ - malignant hyperthermia/Darier disease
K+ - long Q-T syndrome
what are beta-barrel structures
form hydrophobic porins in the outer membrane of bacteria and mitochondria
difference between endo and exonucleases
endo - break nucleic acids at the interior
exo - removes nucleotides from the ends
what type of ligase is most used for preparing plastids
T4-DNA ligase
what % of cells take up the DNA
0.01%
function of tetracyclin and how does it distinguish
acts as a selectable marker to distinguish recombinant and self-ligated vectors.
recombinant if there is no tetracyclin resistance
do bacteria glycosylate proteins
no
