Understanding interactions between macromolecules Flashcards Preview

BS2091 Biochemistry I: from genes to proteins > Understanding interactions between macromolecules > Flashcards

Flashcards in Understanding interactions between macromolecules Deck (47):
1

What do 2 single-stranded DNA molecules make?

Duplex DNA

2

What do DNA and a transcription factor make?

DNA-transcription factor complex

3

What does an enzyme and a substrate make?

An enzyme-substrate complex

4

What does an enzyme and a drug make?

An enzyme-drug complex

5

What does a cell surface signalling receptor and a hormone make?

A hormone-receptor complex

6

What does an antibody and an antigen protein make?

An antibody-antigen complex

7

What is the equation for the dissociation constant?

A + B AB

Kd = ([A] [B]) / ([AB])
Units are concentration

8

What is specificity?

The difference in KD for one site versus another

9

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

10

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.

11

A high degree of specificity does not necessarily mean..

High affinity, and vice versa

12

What methods can you use to measure KD?

Filter binding
Fluorescence anisotropy
Isothermal calorimetry
Equilibrium dialysis
Surface plasmon resonance - BiaCore
Optical interferometry - Octet

13

Two molecules will interact if there is a loss of..

Gibbs free energy on the formation of a complex

14

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

15

Describe enthalpy

H-bonds, electrostatic interactions, van der Waals interactions

16

Does freeing bound water increase or decrease entropy?

Increases entropy

17

Does rigidifying macromolecules increase or decrease entropy?

Decreases entropy

18

What types of interactions can you get in terms of rigid and flexible?

Rigid:rigid
Rigid:flexible
Flexible:flexible
(Don't forget the solvent)

19

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

20

What part of the DNA helix can help to achieve specificity?

The major (wide) and minor (narrow) grooves

21

What charges do the blue, red and white parts of the chemical space-fill models represent?

Blue - positive potential
Red - negative potential
White - neutral

22

What properties does the 434 repressor binding site have?

Reading head
4-6 base pairs
HTH motif
Helix dipole
Dimer

23

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

24

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

25

How many times does a 12 base pair binding site occur in the human genome?

Around 200 times

26

What is the KD value for the 434 repressor?

10^-9 M

27

What properties does the CAP activator binding site have?

HTH motif
Bends DNA

28

What properties does the engrailed homeodomain binding domain have?

Eukaryotic HTH
Not a dimer
Arginines in the minor groove

29

What properties does the met repressor binding domain have?

Beta-strands in the major groove

30

Give examples of zinc-binding DNA recognition motifs

Classical zinc finger (tramtrack), e.g. zif 268 - zinc finger contains tandem repeats to read more sequence
Hormone receptor, e.g. oestrogen receptor DNA binding domain (GATA-I) contains half a NR DBD and a long tail in the minor groove
GAL4
TBP-TATA binding protein contains a protein saddle in the minor groove which is splayed open

31

Are cartoon chemistry figures accurate or misleading?

Misleading

32

What does the CAP activator help to do?

Bend DNA

33

Name a protein that can cause extraordinary deformation of the DNA

TBP-TATA binding protein

34

What are the lifetimes of protein:protein interactions?

Obligate (often co-folding)
Transient (often weak but highly specific)
Intermediate

35

What types of interaction can be protein:protein interactions?

Rigid:rigid
Rigid:flexible
Flexible:flexible

36

What drives the interaction of protein:protein interactions?

Role of enthalpy and entropy

37

What is a rigid:rigid interaction also known as?

Lock and key

38

What is a rigid:flexibility interaction also known as?

Induced folding

39

What is a flexible:flexible interaction also known as?

Co-folding

40

What is the KD value for the barnase:barstar interaction?

10^-14 M (= Koff/Kon)

41

Describe the barnase:barstar interaction

Buried residues at the interface
Importance of charge steering on on rate

42

Can transient interactions also be co-folding?

Yes

43

What do charged surfaces cause?

Electrostatic steering

44

Are affinity, specificity and discrimination the same or different?

Different

45

What is the additive effect of interactions also called?

Multiplies affinity

46

In terms of protein:drug interactions, the affinity of interaction is often driven by entropy, hence often largely..

Non-polar character

47

In terms of protein:drug interactions, for specificity, sometimes shape is enough, but often form precisely..

Positioned hydrogen bonds