enzymes Flashcards
how do enzymes decrease activation energy?
they provide catalytically competent groups for specific reaction mechanisms
by providing substrates such that their orientation is optimised for the reaction
by prefertially binding and stabilising the transition state(s) of the reaction
what are 3 key features of the active site of enzymes
3D space comprising of crucial amino acid residues - provides specifity due to unique 3D arrangment
binds substrate via multiple weak interactions
may only represent a small part of the protein structure
How is enzyme activity measured?
using the michaelis- menton model
what is Vmax?
theoretical maximum rate of reaction - when all enzymes are saturated with substrate
what is Km?
michaelis constant defined as the substrate concentration at half the maximum velocity
what are 4 factors affecting the rate of enzyme catalysed reactions?
substrate/enzyme concentration
temperature
pH
inhibitors
what is reversible inhibition?
inhibition usually non-covalent
ibuprofen - competiitve inhibitor but not covalently attached.
what is irreverisble inhibition?
covalent modification of rthe enzyme usually at AA side chains in the active site
e.g. Aspirin which covalently modifies a serine residue in the active site, it is a competitive inhibitor
what is competitive inhibiton?
either substrate or inhibitor can bind to enzyme but the inhbitor usually binds to the same site as the substrate - the active site , can be overcome by high substrate concentrations
e.g. iBuprofen
what is non competitive inhibition?
inhibitor and substrate can bind simultaneously and binding occurs at a independent sites, it alters the conformation or accesibility of the active site
e.g. methotrexate inhibits dihydrofolate reductase
what happens to Vmax and Km in compeitive inhibiton?
Vmax is unchanged
Km is increased
a higher substrate concentration is needed to reach Km
what happens to Vmax and Km in non-competitive inhibition?
Vmax is decreased
Km is unchanged
increasing substrate concentration has no effect
what are IC50 values?
inhibitor concentration that reduces enzyme activity by 50%
so higher IC50 = lower affinity the inhibitor has to the enzyme so less effective an inhibitor
what are cofactors?
inorganic ions essential for enzyme function
what are the roles of metal ions as cofactors?
metal ions may be part of the active site/ involved in electrostatic substrate binding
metal ions may act as redox agents
metal ions may regulate activity of enzymes
what are coenzymes?
carriers of reaction components, including many water-soluble vitamins
what do NADH and FADH2 do?
carry electrons
what does coenzyme A do?
carry acetyl units
what does bibtin and thiamine pyrophosphate carry?
CO2
What does mutation in Glucose-6-phopshate dehydrogenase cause?
G6DPH deficieny causes metabollic defects
X linked recessive so most carriers are asymptomatic.
G6DPH produces lots of the bodys NADPH (which is needed to drive biosynthesis of nucleic acids and lipids atc)
a crisis can be triggered by certain drugs or food or infection
high incidence in malarial region, it results in ;primaquine induced haemolytic anaemia
what is primaquine?
anti malarial drug
what is favism?
development of haemolytic anaemia upon consumotion of fava beans
what factors are involved in the control of enzyme activity?
inhibition
feedback regulation
covalent modification - protein phosphorylation, enzymes are phophorylated by kinase enzymes
proteolytic activation - when innactive precursor enzymes are made active by proteolysis
describe the role of serine proteases in blood clotting
lysis of blood clots by plasmin.
TPA (tissue plasmin activator) adheres to clot and binds plasminogen - targets activity to the correct place.
this cleaves plasminogen to active plasmin
plasminogen digest clots to small polypeptides
name 3 examples of enzymes with isoenzymes
lactate dehydrogenase (LDH)
cyclo-oxygenase (COX-1 and COX-2)
creatine kinase
what subunits is LDH made from?
M and H, isoenzymes have different combinations of 4 of these subunits
what subunits are creatine kinase made from?
M and B, made of 2 polypeptides as a combination of these
diagnostic value of enzymes in liver damage?
measure of paracetamol levels in the plasma
measurement of glucose levels - immobilised enzymes on plastic strips gives quick and semiquantitative results
diagnostic value of enzymes in AMI?
creatine kindase
myoglobin
cardiac troponin
lactate dehydrogenase
how do you measure paracetamol levels?
in the plasma
using bacterial enzyme which breaks paracdetamol down into acetate and p-aminophenol
the p-aminophenol reacts to form a coloured dye product
how do you measure blood glucose?
using immobilised enzymes and coloured reagents on plastic strips
how can enzymes be used to treat thrombosis?
treat with TPA activates plasminogen to form plasmin which breaks down fibrin clot of thrombosis into peptides
how can enzymes be used in cancer therapy
asparagine is not needed in normal cells as asparagine synthetase enzymes are present in these cells but tumour cells are deficient in Asn sythetase, therefore Asn is essential for them.
treating with asparaginase breaks down asparagine so the plasma level of asparagine are lowered and the growth of tumour cells is affected as they have no Asn sythetase to produce more.
