Peptides and Protein Chemistry 1; Amino Acids, Peptides, Proteins.

Question 1

What is the difference in molecular weight for peptides and proteins?

Answer 1

• Peptides: usually less than 50 AA residues linked by peptide bonds
• Proteins: more than 50 AA residues (large proteins can contain thousands)

Question 2

What are the characteristic groups of an amino acid?

Answer 2

• Chiral carbon centre
• Amino group (NH2)
• Carboxyl group (COOH)
• Rando hydrogen (H)
• Characterising R-group side chain

Question 3

How can amino acids be classified?

Answer 3

According to their side chains:

• Charged (positive/negative)
• Polar uncharged (hydroxide/amide group)
• Non-polar/hydrophobic (aliphatic/aromatic)
• ‘Special cases’ w/unique properties (Cys/Gly/Pro)

Question 4

How are peptides defined/is there any overlap with proteins?

Answer 4

• Small molecules
• Usually less than 50 AA residues
• Do not possess well-defined 3D structures
• Distinction between peptides and proteins sometimes unclear, particularly with medium-sized molecules (15-50 residues)

Question 5

What is responsible for the main properties of a peptide bond?

Answer 5

The mesomeric effect of the N lone pair with the electronegative O; resonance giving rise to partial double bond character C = N+.

Question 6

What are the two consequences of resonance stabilisation of the amide group?

Answer 6

• The peptide bond is chemically inert
• The amide group is planar; has double bond character

> Electrons delocalised into π orbital (via P)

Question 7

What are the potential issues with synthetic peptide synthesis (not see in nature) e.g. making a dipeptide?

Answer 7

• There is no way of distinguishing between the two amino/carboxyl groups; 3 other dipeptides formed
• Many amino acids possess side chains with reactive functional groups e.g. NH2 of Lys, COOH of Glu, OH of Ser.

Question 8

What is the solution to the issues of synthetic peptide synthesis?

Answer 8

• Protection of amino acid residues (the NH2, R-side chains and -COOH not desired to be reacted) with protecting groups that prevent unwanted reactions
• Condensation (H2O leaves) with now-protected amino acids form peptide bond
• Removal of protecting groups in mild conditions

Question 9

What is chemical activation of amino acids and why is it necessary?

Answer 9

• Attachment of a leaving group (X instead of -OH) of -COOH to facilitate attack by amino component (of other AA)
• Otherwise at ambient temperatures an amine will form a salt with a carboxylic acid instead; NH3+/COO-

Question 10

What does chemical activation do to the carbon of -COOH to facilitate a reaction with an amino group?

Answer 10

Replacing the -OH for -X (e.g. Cl) means the carboxylic C is more delta positive (Cδ+) thus more likely to react; HX is leaving group instead of H2O in the OG

Question 11

What does chemical activation mean for peptide bond formation?

Answer 11

• Rapid and quantitative reaction
• Under mild conditions
• Avoids side-reactions
• Doesn’t affect chirality
• Produces easily removable co-products (HX)

Question 12

What are good leaving groups to chemically activate an amino acid (replace the -OH) with?

Answer 12

Conjugate base of a strong acid (e.g. HCl w/o the H):

• acyl halides
• active esters
• carbodiimide reagents

Question 13

What comprises the backbone of a protein?

Answer 13

The primary amino acid sequence; the peptide bonds and C-alpha atoms.

Question 14

What is a torsion angle?

Answer 14

The angle between the plane of atoms of stuff

Question 15

What two torsion angles define the conformation of the peptide chain and why?

Answer 15

• Angle about the Cα-N bond; phi φ
• Angle about the C α-C bond; psi ψ
• Peptide bond has partial double bond character thus is planar; main chain rotations are thus restricted to the above two torsion angles.

Question 16

How do steric factors influence the conformation of the peptide chain; is there one preferred?

Answer 16

• Cis conformation is unfavourable (e.g. R-groups on the same side = bulky and interfere)
• Thus most peptide bonds are of the trans conformation; R-groups etc. on different sides

Question 17

What is hydrogen bonding and what role does it play in protein structure?

Answer 17

• Hydrogen bound to electronegative atom (e.g. N, O, F) experience electrostatic attraction with another electronegative atom
• Key to formation of secondary structure; alpha helices and beta sheets

Question 18

How does hydrogen bonding feature in alpha helices?

How does it contribute to stability?

Answer 18

• Oxygen of C = O (carbonyl) of residue ‘i’ forms H-bond with H of amide residue ‘i+4’; there are 3.6 amino acid residues per turn of the helix
• Relatively weak in isolation, but summation of H-bonds in helix = stable

Question 19

How are the amino acid side chains arranged in an alpha helices?

Answer 19

They point outwards and towards the N-terminus of the chain; ‘up like a tree’.

Question 20

How does hydrogen bonding feature in beta-pleated sheets?

Answer 20

• C=O and N-H groups form H-bonds between neighbouring strands; may be parallel or antiparallel
• Sheets pleat to maintain correct H-bond stereochemistry

Question 21

How are the amino acid side chains arranged in a beta pleated sheet?

Answer 21

Side chains point alternatatively on opposite sides of the sheet

Question 22

What are β-turns in relation to proteins?

Answer 22

β-turns allow the protein backbone to make abrupt turns; more rare.

Question 23

When is it possible that the same protein/primary sequence can adopt different structures?

Answer 23

E.g. in Mad Cow Disease; native proteins re-fold into a different stable conformation, with fatal consequences.

Question 24

What is Glycine’s unique property as an AA and how does this influence protein structure?

Answer 24

• Simply hydrogen as side chain R; thus no chiral centre either
• Lack of bulky side chain (R) means less steric hindrance; Gly renders a polypeptide more flexible

Question 25

What is Proline’s unique property as an AA and how does this influence protein structure?

Answer 25

• Forms a cyclical secondary amine w/side chain

- The tertiary amide bond formation means that proline restricts the torsion angles in a polypeptide chain

Question 26

What is Cysteine unique property as an AA and how does it influence protein structure?

Answer 26

• Thiol group (R-SH)

- Disulfide bond formation (covalent-type) can occur within same polypeptide chain or between different chains (of Cys)

Question 27

Where proteins w/disulfide bridges normally found?

Answer 27

Often seen in proteins excreted into the extracellular environment; intracellular environment is reducing and therefore these proteins due not generally contain disulfide bridges

Question 28

Define: structural motif

Answer 28

Groupings of secondary structure elements e.g. beta-hairpin (antiparallel strands connected by tight turns)

Question 29

Define: protein domains

Answer 29

Globular-type units that are connected by linker regions, often fulfilling different roles and bind to different binding parameters (bringing together different functionalities)