Tertiary and Quaternary Structure

what is tertiary structure

• its the 3D structure taking into account interactions between back bones and R groups of amino acids

• determined by primary structure

what is Quaternary Structure

• Its the spatial arrangement of each of a proteins sub-units

• Sub units are different polypeptide chains

Bonds that stabilize T and Q structure

1) Disulphide bonds → between cysteine residues, common on extracellular proteins

2) Ionic interactions / salt bridges → between oppositely charged amino acids

3) Hydrogen bonds → between side chains, between peptide groups , between side chain and peptide group and with water

4) Hydrophobic interactions and VDW forces

listed form strongest to weakest

The hydrophobic effect

• helps proteins get into there TS

• Hydrophobic effect helps limit non polor and hydrophobic molecules intercation with water

• this is because the increase in entropy of water would have formed cages aroudn the molecule

• the hydrophobic effect causes chain that has NP hydrophobic side cahins to fold and end up on inside of proetin

• and side chain that are polar and hydrophilic end up on the outside

Hydration shell

• water can interact with polar hydrophilic side chains through H bonds to stabilize protein → called hydration shell

• water can also directly interact with backbone and side chain on the interior protein though buried water molecules

what other compounds can stabilize TS and give an example

metal ions and ligands

e.g. Zn fingers

How do Zn fingers work

they are nucleic acid binding proetins

• finger like strucure includes alpha helix and 2 short antiparalle B sheets held togetehr by Zn

• stabilized by a Zn ion that is tetrahedrally coordinated using CYs,HIs and Asp or Glu side chains

• Zn fingers too small to be stable without Zn

what data base allows for proetin structure to be analysed

Proetin data bank (PDB)

what has been described to be within protein folds

super secondary structure or motifs

• are an intermediate level of organisation between secondary structure and tertiary structure

Examples of motifs

• β-α-β motifs

• Greek keys

• α-α hairpins

• Zn finger motifs (same as before)

• β-α-β motifs

• antiparalle B - sheets are joined by short terns

• parallel B -sheets are joined together with longer turns usually with an alpha helicase segment

• hydrophobic residues interact between alpha and b- sheets

Greek motif

• larger motif

• when have consecutive 3 anitparallel B-sheets connected by small hairpins followed by a larger connection to the 4 B-sheets which lies adjacent to the first

α-α hairpins

• the connection between 2 antiparallel a helicase

• the longer the loop the greater the number of combinations

Domains in pyruvate kinase

has 3 domains:

• a β-sheet domain

  • two are of the α/β type

Quaternary structure describes a protein composed of two or more polypeptides, called

subunits

what are homodimers

multiple of the same sub unit

what are hetrodimers

dimers made up of different sub-units

Collagen

Fibrous protein - structural role

Hemoglobin

Is a Globular proteins - functional role

insulin receptor

type of membrane protein

what are protein dynamics

the movements and conformational changes that proteins undergo to perform their functions

covers a large range of movement amplitudes and timescales

Protein dynamics can be versatile and covers a large range of movement amplitudes and timescale

Sub-angstrom vibrations of covalent bonds represent the fastest of those movements (fs)

Then come side-chain rotations (ps–ns)

and backbone fluctuations (ns),

followed by loop motions (ns–ms),

ligand binding/unbinding events (>100 ns),

and collective domain movement (>µs).

loop motions

when the substrate binds the triosephosphate isomerase enzyme, a loop changes from an ‘open’ conformation (green) to a ‘closed’ one (red), which prevents solvent access to the active site and stabilises the intermediate compound of the reaction

Domain motions

For example, bacterial malonyl coenzyme A (CoA) synthetase undergoes a conformational change when the substrate binds: the C terminal lobe rotates upon ATP / Mg2+ binding

what are Intrinsically Disordered Proteins (IDPs)

proteins that are partially or completely unstructured

IDPs are malleable, adapting to structurally different partners, making them ideal “hub proteins”! IDPs are also thought to be involved many human diseases, including cancer, Alzheimer’s, and Parkinson’s disease

Receptor and ligand associate via

non covalent interactions

such as hydrogen bonding, electrostatic interactions, hydrophobic and Van der Waals forces, and shape complementarity

what does binding affinity describe

strength of binding between receptor and ligand

what is it measured in

equilibrium dissociation constant (Kd),

the reverse of the equilibrium association constant (Ka)

equation for Kd and Ka

The smaller the Kd value …

the higher the binding affinity is, the faster ligand will binds and longer will stay in that conformation

the larger the Kd value ….

the smaller the binding affinity the longer it will take to bind and the fats the interaction will be complex will not last for long

why is knowing drug affinity important

important in drug design the larger the drug affinity is the longer it will bind to target and so only need a small dose and so limiting amount of side effects

most common way to determine the Kd value for a given receptor-ligand complex

vary concentration of Ligand in a fixed lowconcentration of receptor

At each concentration of ligand ([L]), the fraction of bound receptor or occupied binding sites (Θ) – is measured

Θ ranges form

0-1

when do we get Kd

when Θ is ½ then, [L] = Kd

So Kd is

Kd can therefore be interpreted as the ligand concentration that leads to 50% occupancy of the receptor’s binding site

Typical Kd values range from

10-15 M to 10-3 M

Chaperones

interact with non- native proteins (unfolded / partially folded or improperly folded proteins)

Assist protein folding by preventing non specific aggregation between protein

or provide microenvironemnt proteins

misfolding

have hydriphobic amino asids on the cell surface which can bind with other molecules causing aggregates

protein aggregation can lead to diseases like; sickle cell anemia, alshizmers, parkinsons , huntintons

Denaturing conditions include:

• Temp

• pH

• chaotropic agents guanidinium ion and urea

• Detergents

• reducing agents

• mechanical stress

Denaturing curve shows

relatively sharp transition from the folded, or native, form to the unfolded, or denatured, form in a sigmoid curve

half way in the curve

find temperature Tm where 50% folded and 50% unfolded

Kd= 1

from here we can determine ..

denaturant concentration at which the protein is 50% unfolded (C1/2).

We determine the temperature at which the protein is 50% unfolded (Tm or melting temperature)

K_eq = 1

ΔG⁰ = 0