Understanding interactions between macromolecules Flashcards
(47 cards)
What do 2 single-stranded DNA molecules make?
Duplex DNA
What do DNA and a transcription factor make?
DNA-transcription factor complex
What does an enzyme and a substrate make?
An enzyme-substrate complex
What does an enzyme and a drug make?
An enzyme-drug complex
What does a cell surface signalling receptor and a hormone make?
A hormone-receptor complex
What does an antibody and an antigen protein make?
An antibody-antigen complex
What is the equation for the dissociation constant?
A + B AB
Kd = ([A] [B]) / ([AB])
Units are concentration
What is specificity?
The difference in KD for one site versus another
Why is specificity important?
For proteins: There are many families of similar proteins For DNA: Binding sites in a sea of other DNA For drugs: On- and off- target activities
In reality there is a continuum of different binding affinities..
And a particular protein will be distributed among binding sites according to the different KD’s.
A high degree of specificity does not necessarily mean..
High affinity, and vice versa
What methods can you use to measure KD?
Filter binding Fluorescence anisotropy Isothermal calorimetry Equilibrium dialysis Surface plasmon resonance - BiaCore Optical interferometry - Octet
Two molecules will interact if there is a loss of..
Gibbs free energy on the formation of a complex
What is the equation for Gibbs free energy?
(Constant temperature)
deltaG = deltaH - (T deltaS)
where deltaH is enthalpy (heat), and deltaS is entropy (disorder)
Both require intimate contact between two molecules
Describe enthalpy
H-bonds, electrostatic interactions, van der Waals interactions
Does freeing bound water increase or decrease entropy?
Increases entropy
Does rigidifying macromolecules increase or decrease entropy?
Decreases entropy
What types of interactions can you get in terms of rigid and flexible?
Rigid:rigid
Rigid:flexible
Flexible:flexible
(Don’t forget the solvent)
How can you measure the change in enthalpy and the change in entropy?
Measure KD’s at several different temperatures
Using isothermal calorimetry
delta G = delta H - (T delta S) = RT ln Kd
What part of the DNA helix can help to achieve specificity?
The major (wide) and minor (narrow) grooves
What charges do the blue, red and white parts of the chemical space-fill models represent?
Blue - positive potential
Red - negative potential
White - neutral
What properties does the 434 repressor binding site have?
Reading head 4-6 base pairs HTH motif Helix dipole Dimer
Do hydrogen bonds have a strong or weak dipole? And how does this affect alpha helices?
Hydrogen bonds have a weak dipole
Alpha helices ‘add up’ weak dipoles
Is a 6 base pair binding site specific enough? And explain why
The human genome is ~3 billion base pairs
If sequence is random then a 6 base pair binding site will occur >500000 times
Therefore 6 base pairs is not enough to be specific
