3d structure of proteins

Question 1

what is functionality dependent on ?

Answer 1

• Functionality is dependent upon the 3d arrangement of the polypeptide chain

Question 2

what is the folding of a polypeptide chain determined by?

Answer 2

o The amino acid sequences which is determined by :
o The molecular structure which obtain its properties through the composition of the variable side chain and properties of its amino acids.
o It is also influenced by The molecular environment (ligands, cofactors, solvents and salts)

Question 3

what are properties of amino acids determined by?

Answer 3

• Properties of amino acids are determined by their structure and amino acid composition and fall into categories defined by
o Structural and chemical properties

Question 4

recall charged amino acids.

Answer 4

• Charged amino acids – asp, glu, lys,arg are charged because of the charged groups in the variable side chains
o May be subdivided into acidic (negatively charged)or basic (positively charged)
o Aspartic acid and glutamic acid are negatively charged due to a carboxyl group in their side chains whilst lysine and arginine are positively charged due to an amine group in their side chains.
o Charged amino acids are also polar in nature but are sufficiently different from the polar amino acids to be in the group of their own.

Question 5

recall nonpolar/hydrophobic amino acids

Answer 5

• Non-polar / hydrophobic amino acids – ala, val, phe, pro, met, isoleucine, leucine
o Can be subdivided into aromatic hydrophobic amino acids and non-aromatic hydrophobic amino acids, e.g based on the aromatic nature of the sidechain. Ie into aromatic and nonaromatic hydrophobic amino acids
o This group also contains Sometimes the sulphur containing amino acids are a separate group ; methionine and cysteine are in 2 different groups and contain sulfur, which means in some classifications they are set aside from their original groups into a group called sulfur containing amino acids. However their properties are more accurately described as non-polar and polar aa.

Question 6

recal polar amino acids

Answer 6

• The other group is polar amino acids – ser, thr, tyr, his, cysteine, asparagine and glutamine.
o Tyrosine, histidine and tryptophan are in a subgroup pf polar aromatic amino acid.
o Glycine is sometimes considered nonpolar sometimes it’s a separate group. Glycine only has 1 isoform as its side group is a H, its small and can be found in turns or loops because it provides more freedom of movement
o Histidine is sometimes also considered as a charged amino acid.
o Cysteine again is a special case due to sulfur

Question 7

what is they difference between the polar and charged amino acids ?

Answer 7

• Key difference between the polar and charged amino acids it that they contain oxygen atoms as part of carbonyl groups or nitrogen as secondary amine groups. Ie they contain nitrogen and oxygen as part of carbonyl or amide groups, rather than carboxyl or amine groups.

Question 8

how can we predict the grouping of amino acids ?

Answer 8

–Carboxyl groups COO- -> charged or acidic
–Amine groups NH3+ -> charged or basic
–2o Amine NH and Carbonyl C=O groups ->polar
–Hydroxyl OH -> polar
–Hydrocarbon -> Non-polar or Hydrophobic

Question 9

what determines the structural behaviour of a polypeptide

Answer 9

•The structural behaviour of a polypeptide chain is conferred by the chemical composition of the constituent amino acids

Question 10

what determines the physiochemical properties of the chain and how it folds?

Answer 10

o Properties of the polypeptide changes as you move along the chain and the individual residues confers those properties on the part of the chain and this determines the physiochemical properties of the chain and how it folds.

polypeptide backbone itself contains amide and carbonyl groups which are polar and these play a role in hydrogen bonding forming secondary structure

Question 11

describe the peptide bond

Answer 11

• The peptide bond is a flat planar structure
• the peptide bond imparts restrictions on the folding of the chain
• The peptide bond is quite unusual in that it is a flat planar structure that has a fixed arrangement. This is as a result of delocalised electrons associated with the carbonyl group and amide groups. This means that the peptide bond behaves as if it were a double bond but is not.
• However the rest of the chain has rotational freedom around the bonds of the c alpha atom ie the c alpha- c prime and c alpha and the nitrogen.
• The combination of provides rotational freedom around one set of bonds but not the other.
• And this provides the freedom of movement to dynamically form the complexity of secondary, tertiary and quaternary structure

Question 12

what favours the formation of specific structures

Answer 12

• Rotational freedom of the bonds allows huge variation in the conformation of the peptide chain
• This freedom of movement allows the dynamic nature of proteins and favours the formation of structural arrangements, secondary, tertiary, quaternary structures.

• Amino acids, particulary with large or charged side chains may restrict the rotational movement of the chain.\

Question 13

why is glycine a special amino acid ?

Answer 13

• Glycine is a special amino acid in that the R group consists of a single hydrogen which allows a greater flexibility of the peptide backbone

Question 14

polypeptides adopt a structure based on?—- what does this mean?

Answer 14

energy minimisation

• All molecules have an intrinsic/specific energetic state which determines its conformation. Minimisation of this drives changes towards the minimum energetic state.
• This is determined by the attraction and repulsion forces.
• The energetic state of a molecule is described by gibbs free energy
• The minimisation of this energetic state (the free energy of a molecule “G” – gibbs free energy) determines the most favourable arrangement of the atoms (conformation)
• the change in free energy upon folding is called ∆G
• If ∆G is negative then the molecule will spontaneously fold to form that resting structure.

Question 15

what is the free energy of any conformation affected by?

Answer 15

the molecular environment:

• Aqueous or lipid (a membrane)
• Other proteins or molecules including salts and their ionic state
• Changes in this environment can induce a further conformational change leading to a different structure. Therefore protein structure is dynamic in nature. , eg association with cofactors or binding a ligand.

Question 16

what determines structural stability ?

Answer 16

bonds

Question 17

what is the difference between covalent and noncovalent bonds?

Answer 17

• Main difference is the strength between the 2
• Weak non-covalent bonds have 1/20th strength of covalent bonds.
• But the overall contribution of non-covalent bonds is significant because non-covalent bonds are far larger in number

Question 18

describe the 4 types of non-covalent bonds

Answer 18

1.Charge or electrostatic attractions
•falls off exponentially as distance increases, affected by electrostatic environment (aqueous environment
- Attractions between the opposite charge e.g carboxyl and amino groups of aspartic acid and lysine

2. hydrogen bonds (transient bonds similar in some respects to covalent bonds)
   - hydrogen bonds occur because of a shared hydrogen between polar groups like the carbonyl and amide groups

• 3. van der waals attractions or dipole interactions are weak forces that occur between two atoms that result from the fluctuating charge associated with the electron cloud
• and is an equilibrium between opposing forces between two atoms determined by the van der waals radius of the atoms involved
• 4. hydrophobic interactions form the fourth weak force and can be thought of as the minimise disruption of water network
• water of course is a polar molecule that forms a network of polar interactions and a lattice network that is in a dynamic equilibrium, thus hydrophobic interactions are driven by the minimisation of the disruption of the water lattice
• And thus a protein will adopt a structure that minimises that network
• for example forming a globular structure with hydrophilic groups on its external surface in an aqueous environment

Question 19

covalents bonds in polypeptides ?

Answer 19

Covalent bonds involved in the formation and stabilization of a folded structure other than the covalent bonds of the polypeptide back bone consist of disulphide bonds between neighboring cysteines

Question 20

how and where do disulphide bonds form?

Answer 20

Disulphide bonds form between neighbouring cysteines of the side chains of two cysteine residues

• The SH groups from each cysteine cross link
• Usually occurs in distant parts of the primary sequence but adjacent in the 3d structure.
• Can form on the same (intra-chain- tertiary) or different (inter-chain-quaternary) polypeptide chains, eg insulin
• They form because of an oxidative reaction between the two cysteines that come into proximity as a consequence of folding.

Question 21

why does folding of the polypeptide occur?

Answer 21

Folding occurs in order to minimise the energy of a molecule and form the structure with the lowest free energy

Question 22

what determines the final structure of a protein

Answer 22

• The minimum energetic state

Question 23

what can the change in structure of a misfolded protein lead to ?

Answer 23

• Firstly, the change in the structure of a misfolded protein leads to the function of the mis-folded protein is almost always lost or reduced
• Secondly mis-folded proteins often have a tendency to self-associate and form aggregates

Question 24

what can lead to the loss of protein

Answer 24

• Misfolded proteins are often detected by the cellular machinery and targeted for destruction a good example of this is in cystic fibrosis and loss of the protein as a consequence contributes to the disease

Question 25

how and why do proteins become misfolded?

Answer 25

1. somatic mutations in the gene sequence leading to the production of a protein unable to adopt the native folding
2. errors in transcription or translation leading to the production of modified proteins unable to properly fold
3. failure of the folding machinery- eg chaperones. If the cell undergoes an environmental change such as heatshock, some proteins denatured and recovery of the protein by the chaperone ensues. Failure of the folding machinery results in an accumulation of misfolded protein.
4. mistakes of the post-translational modifications or in trafficking of proteins
5. structural modification produced by environmental changes
6. induction protein mis-folding by seeding and cross-seeding by other proteins

Question 26

recall the amyloid hypothesis

Answer 26

• In Alzheimer’s disease proteolytic cleavage of Amyloid Precursor Protein (APP) is observed
• Beta amyloid is a small protein released as a result of proteolysis from a larger transmembrane protein called amyloid precursor protein, APP.
• APP has multiple functions but is involved in G-protein signalling and is normally anchored in the membrane.
• Cleavage of APP results in a 40 residue peptide beta amyloid is released. This part of the protein is normally lying within the membrane.
• In the intact molecule this anchors the protein in the membrane . APP accumulates in isolation to the rest of the protein it mis-folds to form beta sheets. The beta sheets in turn stack one over the other.

Question 27

outline the steps in aggregation as seen in alzheimers

Answer 27

1) Flat planar monomeric sheets form. Those in turn aggregate to form stacked sheets called oligomers
2) The side chains of the beta sheet reach out from the surface of the sheet. They interdigitate with those of another sheet.
3) The oligomers extend to form protofibrils that further aggregate to form amyloid fibrils
4) The aggregation of beta amyloid in the brain of alzheimers patients is a feature of the pathology of the disease. These interfere with the workings of the synapse, particularly in the hippocampus.
5) Gradually higher order insoluble aggregates form and are deposited in plaques. These damage the neuronal cells of the brain.
6) In the case of beta amyloid, it is an example of post translational modification of the protein resulting in proteolysis leading to diseased pathology

Question 28

describe cystic fibrosis

Answer 28

•In Cystic fibrosis the most common mutation is a single codon deletion of Phenylalanine at residue 508 of the cystic fibrosis transmembrane conductance regulator (CFTR)
The protein regulates chloride ions across the cellular membrane.
- It is a Somatic mutation that leads to the trafficking of the protein that is impaired but still functional to the proteosome and degradation of it occurs there.
•DF508del leads to mis-folding of the protein whilst it is still in the ER
•This is recognised by the cellular mechanism to degrade misfolded proteins via firstly ubiquitination, then secondly traficking to the proteasome and degradation of the protein.
Consequently, very little traffic through the normal route and end up in the membrane.

Question 29

describe induced protein misfolding

Answer 29

The alpha helices are misfolded and form beta sheets .
•Mis-folded proteins (PrPSC)that interact with other normal proteins (PrPC)
•Through this interaction they induce mis-folding of the normal protein and polymerisation. Monomors induce misfolding and aggregation forming oligomers
•Oligomers form fibrils and plaques of mis-folded protein
• key to the stable formation of oligomers is the change in the energy that represents a lower energetic state in the oligomeric structure than in the correctly folded molecule in isolation. Induction in this way, by seeding and cross seeding by misfolded proteins such as prion proteins are examples of protein folding as a dynamic process. This process is reliant upon the concept of energy minimisation ∆G
•It is a dynamic process brought about by the interaction of molecules resulting in a more stable aggregated structure

Question 30

what are prions ?

Answer 30

Prions: Prions are misfolded proteins with the ability to transmit their misfolded shape onto normal variants of the same protein