BIOCHEMISTRY Flashcards
what are proteins
complex biopolymers whos structure is determined by the sequence of amino acids
how many amino acids are there and how are they different
20, differentiated by the R side chains
describe the structure of an amino acid (left to right)
starts with and amine group (NH3) then the alpha C with H ontop and R group below then there is the carboxyl group (COOH)
why are amino acids called zwitter ions
they both a negatively charged end (carboxyl group) and a positively charged end (amine group)
how many stereoisomers exist
2 steroisomers called enantiomers/optical isomers
which enantiomers are found in living things
L-animo acid enantiomers. it is asymmetric and this allows for highly specific molecule recognition
what are the different groups of amino acids
- nonpolar and hydrophobic
- aromatic
- polar,hydrophillic and acidic
- polar,hydrophilic and basic
- hydroxylic, sulfur containing and amidic
why is methionine special
conatins sulfur and is the first amino acid in a polypeptide chain
what is special about cyteine
contains sulfur and is highly reactive and therefore used in enzyme active sites. it can also form disulfide bonds
how does a peptide bond form
when the OH from a carboxyl group links to the H from the amine group they from water so the (C=O-NH) is the peptide bond
what does the primary structure mean
the primary structure determines how the peptide folds into a 3D shape
what is the secondary structure
it is how it is more complexly folded eg alpha helix, beta pleated sheets
what type of bond is the secondary structure dependent on
on hydrogen bonds between O and H
what does it mean that the alpha helix is right handed
the side chains are pointing outwards from the central helix
which direction can the beta sheets form
either parallel or anti parallel
what is the tertiary structure
this is the arrangement of the secondary structures forming a more complex molecule
what are the different types of bonds in the tertiary structure
electrostatic attraction, hydrogen bonds and van der waals forces
what is the quaternary structure
the 3D structure which is several subunits bonded by non covalent bonds
what are the functions of proteins
structural proteins catalytic proteins-enzymes signalling proteins proteins involved in cell adhesion and recognition membrane transport proteins
examples of structural proteins
- extracellular matrix proteins eg collagen and elastin
- muscle proteins
- cytoskeletal proteins
- antibodies, complements
how to detect proteins
- using SDS page gels which are 1D and 2D
- mass spectrometry
- crystallography
- edman sequencing
what is the function of ubiquitin
- regulates major cellular processes such as cell division, immune responses and embryonic development.
- marks proteins for degradation
what is the function of the proteasome in protein breakdown
the proteasome recognizes the ubiquitin and cleaves off the ubiquitin. it then feeds the protein through a channel and then chops it into smaller pieces
how many heme groups does the quaternary structure of haemoglobin have
4
how does abnormal primary structure affect protein function and give examples
- mutations leading to misfolding, sickle cell disease
- mutations leading to trapping of protein in the ER, cystic fibrosis
- mutations leading to premature stop codons and hence unfinished proteins, Duchene Muscular Dystrophy (DMD)
how does abnormal secondary structure affect protein function and which disease does it lead to
leads to amyloidosis which is the collection of amyloid which is a collective name for the congregation of abnormal proteins that cannot easily be broken down. theyre folded into beta sheets leading to extracellular deposits. alot of beta sheets leads to alzheimers disease
what are prions
it is also caused by abnormal secondary structure. prions are proteinaceous infectious particles (no DNA/RNA).
what is the function of a prion protein (PrPC)
normal neuronal protein involved in cell adhesion, ion channel activity and neural excitability
what is the shape of PrPC
normally an alpha helix secondary structure but when abnormal it changes into beta sheets
when ribosomes synthesize proteins which end do they start from
N-terminal of the amino acid, amine side
what type of amino acids can form hydrogen bonds
hydrophillic
which protein detection methods is most suitable to analyse protein structures
crstalography
which protein detection methods is most suitable to check the occurrence of a specific protein
immunoblots
what are enzymes
proteins with unique structures that are folded to create active sites which enable recognition of specific substrates they transform
what do oxidoreductases do
oxidation and reduction reaction
what do transferases do
transfer amino,carboxyl,acyl,carbonyl,methyl,phosphate and other groups between molecules
what do hydrolases do
cleavage bonds by adding water
what do lysases do
cleavage of carbon-carbon,carbon-sulphur and carbon-nitrogen but not peptide bonds
what do isomerases do
rearrangement of bonds
what do ligases do
formation of bonds between carbon and oxygen, sulphur and nitrogen
what is a catalyst
a protein which increases the rate of reaction without beinf changed itself in the overall process
how does a catalyst affect a reaction
helps change the rate not the equilibrium however equilibrium is reached faster
what is the induced fit model
when two substrates fit into the active site and form a enzyme-substrate complex. the activation energy is lowered and the two substrates form one product
what is the transition state theory
when one substrate binds to the active site forming an enzyme substrate complex, it undergoes catalysis and the two products stay in the active site now called an enzyme product complex/transition state and the product is then released
how do enzymes reduce the activation energy
they bind to the transition structure stabilizing it which leads to a lower activation energy
how is the transition state stabilized
the substrate binds in the correct orientation and binds tightly
what is an example of the transition state theory
lysozymes
how does the active site in the induced fit model reduce the activation energy
they bind the substrate in a favorable orientation but also allows the shape to be closer to that of the transition state
what do the enzymes do in the induced fit model
they can change their shape based on the substrate and need a chain reaction to return back to the original shape
what affects the rate of reaction
temperature, pH and substrate concentration
describe a graph of rate of reaction (y )against substrate concentration(x)
it is a curve starting from zero zero and plateauing at the higher substrate concentration.
how can you determine the enzyme capacity by using the curve
the height of the curve is the enzyme capacity
why does the rate of reaction plateau
all the active sites are occupied so that is the maximum rate of reaction despite increasing substrate concnetration
what is the symbol for when the enzymes are fully saturated
Vmax and it is dependent on enzyme concentration. its at the top when it starts to plateau
what is Km (its a constant )
it is the affinity of the substrate for the enzyme. Km is equal to the substrate concentration at which the enzyme converts substartes into products at half of its maximal rate
what is the relationship between the Km and the affinity for the enzyme
the higher the Km the lower the substrates affinity for the enzyme
what does having the same Vmax and having the same Km mean
same Vmax means- same enzyme concentration
same Km means - same affinity for the enzyme
what are isoenzymes
catalyse the same reaction but different structures
what is cytochrome P450
- catalyse oxidation
- superfamily of microsomal (liver) enzymes
- named as CYP+number+letter
what are competetive inhibitors
- they bind reversibly to the active site
- Km increases (reduces affinity for the enzyme)
- Vmax remains unchanged
what are non-competetive inhibitors
- inhibitor binds outside the active site, decreasing raate of reaction
- Km remains unchanged
- Vmax decreases
what are allosteric regulators
-they are multi-subunit proteins with multiple active sites
what happens during allosteric regulation
-inhibitors bind to a site that is not the active site and this reduces Vmax and increases Km(reduces affinity)
what is an allosteric activator
attaches outside the active site and it allows the substrate to bind at a higher affinity
what is feed forward activation
when a metabolite produced activates an enzyme that catalyze a reaction further down the pathway
give an example of feed forward activation
pyruvate kinase in glycolysis is activated by fructose-1,6- biphosphate
what is gibbs free energy
the energy of products minus the energy of the substrate
what is the purpose of biochemical profiling
to assess the physiological status of an individual
what is patient stratification
separating patients to different treatment streams based on some characteristic
how to measure biomolecules in the laboratory
each molecule absorbs light differently and at a certain wavelength. if a light source passes through a solution containing those molecules the amount of light absorbed will be in direct proportion to the number of molecules
what equipment is used and what are the units
spectrophotometer in optical density (OD)
what are colorimetric assays
this is when biomolecules being measured produce a color complex of a particular wavelength
how is blood glucose measured
the glucose is converted by glucose oxidase into D-gluconic acid. simultaneously the glucose oxidase coverts the oxygen into hydrogen peroxide which reacts with horse radish peroxidase (HRP) which forms a color complex with a wavelength absorption of 540-570nm
how do you convert optical density values into concentrations
the optical density of known concentrations of the sample are measured and plotted. a standard curve or line of best fit is drawn and the concentration can be found using the OD of the unknown samples
what are screening tests
provide preliminary physiological data to reflect the general conditions of an individual. less complicated tests are involved and normally can be performed within a short period of time
what are specific function tests
will provide more details to indicate whether a particular body system is performing their normal function or if there is any damage to particular organs
what is the most common biochemical test request
urea and electrolytes
what is the difference between blood plasma and blood serum
plasma is the liquid, cell free part of blood that has been treated with anticoagulants whereas serum is the liquid after coagulation therefore void of fibrinogen and other clotting factors
why and how do we use serum as a test
whole blood is collected and the samples will be left to allow for clot formation and all the clot with blood cells is removed by centrifugation since fibrin and cellular components interfere with assay results
why and how do we use plasma as a test
specific types of tests may require preservation of analyses by addition of enzyme inhibitors or preservatives e.g. adding fluoride oxalate to prevent the breakdown of the glucose being tested
why is venous blood sampling more common than arterial
due to high pressure in arteries are compared to veins
when do we use venous blood samples
for analyzing blood cell contents
- glycosylated hemoglobin( raised level in diabetes )
- white blood cell count
- red blood cell count
when do we use arterial blood samples
- blood gas analysis for arterial pressure of O,CO2 and pH
- blood gas analysis to indicate hypoxaemia
when do we use capillary blood samples
- in young children
- people with fragile veins e.g. old people
- patients with severe burns
how can we manage acid base disorders
if symptoms are indicative of an acid base imbalance the first step is to collect arterial blood and examine the hydrogen ion concentration. then we consider change in pCO2 and HCO3 and manage accordingly
why do we look at HCO3
it forms an important part of the buffering systems in our body
how is urea made
urea is the breakdown product of amino acids processed to ammonia in liver. ammonia plus CO2 is processed to urea
how do we measure urea in clinical samples
-take a sample of clotted venous blood from your patient and extract the serum after centrifugation. urease converts urea to ammonium. 2-oxyglutamate and ammonium is converted to glutamate using glutamate dehydrogenase (GLDH) which also converts NADH to NAD and NADH measured at 340nm
what is the relationship between NADH and urea concentration
the higher the absorbance of NADH the lower the concentration of urea
what key roles do electrolytes undertake
osmolarity, pH maintenance and biochemical reaction co enzymes
what are three rules when interpreting electrolytes
- one third of the fluid is extracellular where as two thirds is intracellular
- water isn’t restricted to one compartment but sodium is
- water follows sodium so if sodium leaves water leaves leading to dehydration
which ions are present more in the ICF than the ECF
potassium
magnesium
phosphate
which ions are equally present in the ECF and ICF
calcium
which ions are more in the ECF than the ICF
sodium
chloride
bicarbonate
how do we measure electrolytes
using an ion selective electrode ,ISE
how does ISE work
there are two electrodes , the reference electrode allowing all ion influx into the electrode and sample/indicator electrode which only allows a certain electrode
this creates a potential difference and so the current created reflects the abundance particular ion in the sample
what are possible conditions linked to electrolyte imbalance
- chronic renal malfunction
- dehydration
- liver failure
what is reference range
It indicates the concentration range within which results of a non-symptomatic individual would fall into
what does a liver function test measure
- total protein
- albumin
- bilirubin
- enzymes
what enzymes does a liver test measure
- alanine aminotransferase ALT
- aspartarte aminotransferase AST
- alkaline phosphatase ALP
- gamma glutamyl transferase
what protein is used to analyze heart damage and what do abnormal levels indicate
analyses by cardiac troponin and shows ischaemic cardiac damage
what proteins form the troponin complex
TnC ,TnI and TnT
why is detection of TnI and TnT more specific than TnC
TnC is released by other muscles while the other two are specific to the cardiac muscles
how do we use measures of Tn levels to show heart damage
comparison between time points e.g. between three hours since the start of symptoms and if there is a 20% increase it shows cardiac muscle damage
what is an epitope
a specific part of a protein the antibody binds to
how can we detect troponin in hospitals
magnetic beads are coated in anti cTnI (cTnI means cardiac TnI) antibodies against one epitope
the anti cTnI are conjugated to alkaline phosphatase ALP for a second epitope
this allows for the antigen in the patient serum if present to be sandwiches between the two antibodies and then a chemical is acted which reacts with ALP to produce a light measured by a luminometer
what are the three ways pH is maintained
- buffering of intracellular and extracellular buffers
- alveolar ventilation which controls PaCO2
- renal H+ excretion which controls plasma HCO3 conc
what are the major buffering systems of the body
- HCO3- / H2CO3 buffering in the lungs
- HPO4- / H2PO4- buffering
- NH3 / NH4+ buffering in the kidney
how does the carbon buffering system work
when excess H+ ions are added to bow fluids due to an acid they combine with HCO3- and produce H2CO3 which is broken down into H20 and CO2 which is removed by respiration so the buffer system never reaches equilibrium
what are the three steps of gas transfer in the lungs
- O2 binds to deoxyhaemoglobin and forms oxyhemoglobin with the release of H+
- the H+ binds to HCO3- to form H2CO3
- the H2CO3 dissociates into CO2 and H2O
what are the three steps of gas exchange in tissues ( providing them with oxygen )
- oxygenated Hb dissociates to release O2 into the tissues and the deoxygenated Hb is produced
- CO2 generated in the tissues is hydrated to form H2CO3 which ionizes to form H+ and HCO3-
- the deoxygenated Hb accepts the H+
what is the structure of Hb
four subunits 2 alpha and 2 beta each with haem groups
there Is a ferric ion Fe2+ in the center of the haem group
what happens during oxygen unloading
once the oxygen is unloaded in the tissues 2,3-bisphosphoglycerate (BPG) binds to beta chains
the 2,3 BPG binding induces a confirmational change of all four chains disabling O2 binding so it cant be taken away
when 2,3 BPG levels are low it dissociated from Hb enabling O2 to bind again
how is haemoglobin synthesised
occurs in the mitochondria of an immature RBC
the Fe2+ ions are delivered by transferrin where they translocate to the mitochondria
inside the mitochondria the alpha aminolaevulinic acid ALA is synthesised, processed in the cytoplasm and is transported back to the as protoporphyrin which binds the Fe2+ to form haem groups
what is reparatory acidosis
failure of the lungs to excrete sufficient CO2
what is metabolic acidosis
failure to excrete enough endogenous acid by the kidneys
what 2 functions must the kidney perform to maintain pH
- reabsorption of all filtered HCO3-
- excretion of the daily H+ load
how can sulphuric and phosphoric acid be removed through the carbonate buffer system
H2SO4 + 2NaHCO3- goes to NaSO4 + 2H2CO3–> 2H2O + 2CO2
where does the phosphate come from
high conc in the cel. many processes use ATP and signaling pathways use phosphorylation
where does the ammonia come from
generated from the amino acid glutamine. ammonium ions NH4+ are excreted as a chloride salt NH4Cl
