Enzymes and the principles of catalysis Flashcards
Describe ATP hydrolysis
- reaction far from equilibrium in vivo
- thermodynamically favourable
- spontaneous
- slow: in the absence of a catalyst, it takes many hours for ATP to be hydrolysed to ADP + Pi
- liberates free energy used to power the cell
Describe TPI
- uncatalysed reaction rate: one a day
- reaction rate when catalysed by TPI: 4,300 per
second
TPI
triose phosphate isomerase
Most enzymes accelerate reactions to
millions of times faster than the uncatalysed rate
Describe the synthesis of the pyrimidine nucleotide, UMP
- one of the slowest uncatalyzed reactions
- uncatalysed rate of decarboxylation of orotidine
monophosphate to UMP is 1 reaction per 45 million
years - catalysed by OMP decarboxylase,
reaction occurs at a rate of 39 per second - rate enhancement of 1.4 x 1017-fold
Describe enzyme catalysis - the basics
accelerate the reaction by stabilizing the transition state, reducing the activation energy required to reach it
Describe the transition state
- no longer the substrate but is not yet the product
- least-stable and most-seldom occupied species along the reaction pathway
- highest free energy
Enzymes alter
reaction kinetics, NOT equilibria
Describe enzyme catalysis - the specifics
- accelerate the attainment of equilibria
- do not change the standard equilibrium position of the reaction (Keq)
- at equilibrium, the rate of the forward and reverse reactions are the same, irrespective of the presence of an enzyme
Describe the equilibrium position
a function only of the free-energy difference between reactants and products
Describe active sites
- crevice in the enzyme structure
- key catalytic groups are precisely orientated around the bound substrate
- water usually excluded unless it is a reactant (crevice mainly lined with hydrophobic amino acids)
- some hydrophilic residues are present for substrate binding or catalysis
catalytic groups
amino acid side chains
binding sites
bind and orient substrate(s)
catalytic sites
reduce chemical activation energy
Describe covalent catalysis
- covalent bonds formed or broken
- active site contains a highly reactive group that becomes temporarily covalently attached to the substrate during catalysis
Describe general acid-base catalysis
a molecule other than water plays the role of proton donor or acceptor
Describe metal ion catalysis
metals assist via substrate binding, stabilising negative charges on reaction intermediates to facilitate acidity and enabling oxidation reactions
Describe the enhancing proximity of reactants
bringing two molecules close together enhances transfer of function
Describe chymotrypsin
- catalyses peptide bond hydrolysis
- attacks the unreactive carbonyl using a powerful nucleophile
Describe nucleophile
- a species which is strongly attracted to a region of positive charge in another species
- contains an electron pair available for bonding
- either fully negative ions, or have a strongly partial negative (d-) charge somewhere on the molecule
- ‘excess’ electrons
- propensity to form a new covalent bond with an electrophile, which accepts the bonding electrons
species
ion or a molecule
Describe a region of positive charge
a nucleus
Describe a region of positive charge
a nucleus
Describe the chymotrypsin catalytic triad
- generates nucleophilic serine
- acid/base catalyst
- nucleophile
Describe the chymotrypsin mechanism
- enzyme-substrate
- first tetrahedral (oxyanion hole)
- acyl-enzyme
- second tetrahedral (oxyanion hole)
- enzyme-product
Describe the Michaelis-Menten model
- E+S -> ES -> E+P
k1
E+S -> ES
k2
ES -> E+S
k3
ES -> E+P
Vmax
- [S]/[S] + KM
- kcat[ET]
Describe the kinetic response of enzymes
- changes in substrate concentration means that the system tends towards a steady state
- concentration of metabolites at steady state is dependent on the KM and Vmax of the enzymes in the system
Describe enzyme kinetic parameters
- evolved such that metabolite concentrations at steady state are maintained at relatively low levels (mM – low mM range)
- metabolism does not exhaust the solvency capacity of the cell
