2 - Protein Energetics and Dynamics

Question 1

What is the flexibility of a protein dependent upon?

Answer 1

The nature and number of the intramolecular interactions.

Question 2

How do the maximum radii of covalent and Van der Waals bonds compare?

Answer 2

Covalent bonds can be significantly longer range.

Question 3

What mechanical law do all bonds obey?

Answer 3

Hooke’s Law, F=Kx where K is the force constant.

Question 4

What modification is made to Coulomb’s Law for protein modelling?

Answer 4

Instead of being squared, the radius is multiplied by the distance dependent dielectric of the solvent, εr.
E= (Qa Qb)/(4π ε0 εr r)

Question 5

What three things need to properly understood about a protein for molecular dynamical modelling?

Answer 5

Dynamics
Folding
Energetics

Question 6

What are forcefields in molecular dynamics modelling?

Answer 6

The way in which all particle interactions are combined to derive the effect of each particle on all those surrounding it and vice versa.

Question 7

What are the two main components estimated in molecular dynamic modelling?

Answer 7

Coulombic Interactions and resultant Newtonian Mechanics.

Question 8

What issue arises in terms of molecular dynamic modelling from the nature of the interactions involved?

Answer 8

According to coulombs law and similar, the distance over which the charge exerts a force is theoretically infinite, though decays quickly. Including negligible forces would increase computing time exponentially, so a distance cutoff is used.

Question 9

What are the three types of cutoffs used for long range forces in molecular dynamic modelling?

Answer 9

Truncated cutoffs
Switch cutoffs
Shift cutoffs

Question 10

How is the energy initially distributed between the atoms in molecular dynamic modelling?

Answer 10

Randomly according to the Maxwell-Boltzmann distribution.

Question 11

How is time factored into molecular dynamic modelling?

Answer 11

After the energies of and forces on each atom have been attributed they are allowed to move as they would for a set ‘timestep’, after which all the parameters are recalculated.

Question 12

What is used to smoothen the motion component of molecular dynamic modelling?

Answer 12

Leapfrog algorithms (AKA Verlet algorithms). These syncopate the timesteps providing intermediate values.

Question 13

What issue is caused by the solvent in molecular dynamic modelling?

Answer 13

The huge number of solvent molecules required to simulate in order to create a representative model of a protein mean that water atoms often greatly outnumber the more interesting protein atoms. Calculating the molecular dynamics of the water then takes up most of the computation time.

Question 14

How is the solvent issue in molecular dynamic modelling solved?

Answer 14

Using ‘periodic boundaries’ in which the water for a given point is simulated and repeated in cubes around the protein.

Question 15

What are five ways in which hydrophobicity is measured?

Answer 15

Hydrophobicity chromatography
Partitioning measurements
Accessible surface area measurement
From thermophysical measurements
Lyotropically

Question 16

What four things are hydrophobic burying dependent upon?

Answer 16

Solvent properties
Salt concentration
pH
Temperature

Question 17

What does hydrophobic protein binding require?

Answer 17

Two surface hydrophobic patches that are highly complementary in shape so as to ensure water exclusion.

Question 18

Other that hydrophobic binding, what is the other main method of inter-protein binding interaction?

Answer 18

Charge complementarity.

Question 19

What programs can be used to simulate protein binding?

Answer 19

GRID, DOCK and Autodock

Question 20

How much of the proteome is thought to be mad up of proteins which are totally intrinsically disordered?

Answer 20

> 20%

Question 21

How much of the proteome is proteins with intrinsically disordered regions longer that 30 residues?

Answer 21

50%

Question 22

What type of residue is common to IDPs?

Answer 22

Highly polar ones, preventing any hydrophobic interactions which are the main cause of folding.

Question 23

How many conformations did Levinthal estimate there were through which a protein would need to search through to fold randomly?

Answer 23

10^143

Question 24

What did Anfinsen’s experiment ultimately prove?

Answer 24

That for many proteins the ability to fold is encoded only within the primary structure.

Question 25

What are the two main models of how proteins fold?

Answer 25

Diffusion-Collision

Nucleation Condensation

Question 26

What does the Diffusion Collision model of protein folding posit?

Answer 26

Protein folding begins with hydrophobic collapse into a nucleus around which secondary structure forms before colliding with each other and forming tertiary structure.

Question 27

What does the Nucleation Condensation model of protein folding posit?

Answer 27

Secondary and tertiary structures are made simultaneously.

Question 28

What three imaging techniques are used to investigate protein folding?

Answer 28

NMR, CD, Optical tweezers

Question 29

How do optical tweezers work?

Answer 29

Highly focused lasers exert a photonic radiation pressure called scattering force on a bead, suspending it in the desired position. The force required to move it in a given direction can be measured.

Question 30

How are optical tweezers used to examine protein folding and unfolding?

Answer 30

By attaching a protein to DNA linkers by mutagenic Cys residues on the N and C termini that tie it to a fixed bead and one which can be manipulated by optical tweezers. The force required to pull apart specific folds can then be measured.

Question 31

What general results might be seen from optical tweezer investigation of protein folding?

Answer 31

Abrupt changes in the force required to manually stretch a protein correspond to large unfolding events, one of which will often correspond to the denaturing of stable core of the protein.

Question 32

What are molten globule proteins thought to be?

Answer 32

Either folding intermediates or trapped states.

Question 33

What are the defining characteristics of molten globule proteins?

Answer 33

A high level of dense secondary structure with no defined tertiary structure. They are compact but less so than a nucleation produced core.

Question 34

Where are molten globules found on a folding funnel diagram?

Answer 34

In a horizontal band in the centre of the vertical axis.

Question 35

What is a serious limit to protein modelling?

Answer 35

Inability to simulate protein folding.

Question 36

Why can molecular dynamic modelling not be used to simulate protein folding?

Answer 36

Inaccuracies in forcefield production

Huge amount of conformational space to be explored.

Question 37

What method can allow for highly rudimentary in silico protein folding be guessed at?

Answer 37

Lattice modelling, in which the amino acids are given discrete values (often just polar/non-polar) and folded by the computer into the most favourable chain it can within a cubic lattice.

Question 38

What folding event can be predicted quite well by molecular dynamic modelling?

Answer 38

Unfolding.