proteins

Question 1

primary function of amino acids

Answer 1

monomers which make up the proteins biological polymers.

Question 2

amino acid structure brief

Answer 2

composed of four groups around central carbon atom which is called the alpha carbon (α carbon):

      H H2N-C-COOH
      R

Question 3

what 4 groups do we see in amino acids:

Answer 3

• A variable R group (also called side chain) which is how each unique amino acid is identified. There are 20 different R groups on proteinogenic amino acids
• A hydrogen atom
• An amino group. This is a basic group which is protonated (positively charged) at basic and physiological pH, NH3+.
• A carboxylic acid group. This is a weakly acidic group, which is deprotonated (negatively charged) at physiological pH, COO-

Question 4

• example of a zwitterion (ionised amino acid)

Answer 4

alanine
H
H3N+ - C - COO-
R

Question 5

amino acids are ionisable - what does this mean?

Answer 5

they can be simultaneously positively and negatively charged = producing the zwitterion (a compound that possesses both charges simultaneously, creating a molecule that bears no net charge

→ the pH at which each acid bears net zero charge varies depending on the nature of the R group.

Question 6

why do we use titration curves?

Answer 6

to understand solution ionisations of particular amino acids:

Question 7

https://www.notion.so/proteins-1c800bb3982d80929df2c1bc2b412383?pvs=4#1c800bb3982d80f7ac1cf54562b59e9f

what can we tell from that pH curve:

Answer 7

• At low pH (low equivalents of OH-) the amino acid is fully protonated i.e. the amino acid exists in its cationic form with NH3+ and COOH.
• With increasing pH (increasing equivalents of OH-) the carboxylic acid of the amino acid is deprotonated to give the zwitterionic form with NH3+ and COO-.
• With even higher pH (high equivalents of OH-), eventually the amino group of the amino acid is also deprotonated to give the anionic form with NH2 and COO-.

Throughout the middle region between the two different pKas, the amino acid will become zwitterionic. The exact point at which this happens is different for each amino acid. The midpoint between the two pKas is called the pI (isoelectric point).

Question 8

how can we work out the pI (isoelectric point - the point at which the amino acid has no net charge)

Answer 8

pI = (pKaA + pKaB) /2
The pI can be calculated from the pKa values of the amino acid which are on each side of the zwitterion

Question 9

how do r group characteristics affect the acid?

Answer 9

• Hydrophobic. Hydrophobic R groups form hydrophobic interactions with other amino acids, they don’t form hydrogen bonds or ionic bonds, or interact with water.
• Hydrophilic. Hydrophilic amino acids tend to contain electronegative atoms and so are polarisable. These R groups interact with water, often via hydrogen bonding.
• Acidic. Acidic R groups are ionisable, and tend to exist in anionic form at physiological pH. They therefore often form ionic bonds (also called salt bridges) which are important for protein structure.
• Basic. Basic R groups are also ionisable, and tend to exist in cationic form at physiological pH. As for acidic residues, these amino acids are often involved in ionic bonding.

Question 10

peptides and proteins are polymers: how are amide bonds formed?

Answer 10

by the condensation reaction between the alpha amine of amino acid and alpha carboxylic acid of a second amino acid
- the COOH and NH2 form the CONH

Question 11

why are amides strong covalent bonds?

Answer 11

due to resonance: the central carbon-nitrogen bond of the amide has partial double bond character: this means that amide bonds can only be broken by very harsh chemical conditions (100C and 6M HCl) or by enzymes such as proteases and peptidases

Question 12

amide bonds = cis vs trans conformation:

Answer 12

Cis conformation = both substituents are on the same side of the C-N bond.
Trans conformation = the substituents are on opposite sides of the C-N bond which leads to a more stable structure.

→ Cis peptide bonds in proteins are rare – they are common only for proline due to steric clashes caused with side chains of other amino acids

Question 13

the 3D shape of proteins is determined by what bond characteristics?

Answer 13

peptide bonds

Question 14

why are peptide bonds rigid and planar

Answer 14

the partial double bond character, but other bonds on either side between C-N and C-C can rotate: this allows peptide chains to adopt specific shapes for function

Question 15

levels of protein structure:

Answer 15

1. PRIMARY - The amino acid sequence.
2. SECONDARY - Local 3D secondary structure elements which the peptide chain can fold into e.g. α helix, β sheet. These structures are formed by hydrogen bonding. Proteins often contain multiple different types of secondary structure.
3. TERTIARY - The full 3D tertiary structure of a protein formed by non-covalent bonds and disulfide bonds. Secondary structures are found within the folded tertiary structure of a protein.
4. QUATERNARY - The association of multiple individual folded proteins. (polypeptide subunits)

Question 16

what is the primary structure defined by:

Answer 16

the number and type of amino acids linked together forming the polypeptide chain.

• linear chain of amino acids linked by amide bonds forming a polypeptide
• sequence of amino acids (determines its final shape and function)

Question 17

each end of the polypeptide is usually unbound, leaving what groups on each side?

Answer 17

a free amino group on one end, called the N-terminus, and a free carboxylic acid at the other end, called the C-terminus. The amino acids in a polypeptide chain are numbered starting from the N-terminus, progressing in the same direction as translation i.e. N→C.

Question 18

bond involved in primary structure

Answer 18

Covalent peptide bondsbetween amino acids.

Question 19

secondary structure: defined by

Answer 19

the local 3D structures adopted by polypeptide chains

Question 20

bonds involved in secondary structure

Answer 20

hydrogen bonds (non-covalent)

Question 21

secondary structures are created by hydrogen bonding between…

Answer 21

the carbonyl oxygen of one amide bond and the amine hydrogen in the other

Question 22

traits of hydrogen bonds

Answer 22

• non-covalent interactions (weaker than covalent)
• exist bc partial ionisation of C=O and N-H bonds
• H bond shared between the two atoms

Question 23

hydrogen bonding leads to what 2 diff types of secondary structure

Answer 23

alpha helix and beta sheet

Question 24

what are alpha helices:

Answer 24

a common type of secondary structure characterised by a single helix in which
- all peptide backbone C=O and N-H is involved in hydrogen bonding
- all R groups point towards the outside of the helix

Question 25

what are beta sheets

Answer 25

Beta sheets = 2 or more extended peptide strands hydrogen bond to each other to form a sheet-like structure. - also called pleated sheets because the nature of the planar peptide bonds mean that the sheets of peptide fold like a concertina. Hydrogen bonds form between peptide chains which are next to each other in the sheet: the C=O on one chain will form a hydrogen bond with the N-H on the neighbouring chain. Unlike in α helices, not every amide in the peptide is involved in hydrogen bonding.

Question 26

beta sheets can be parallel or antiparallel - what is the difference?

Answer 26

- In parallel sheets, peptide chains which are hydrogen bonding to each other run in the same direction. - In anti-parallel sheets, the peptide chains run in opposite directions, often linked by a turn (a loop of amino acids).

Question 27

what is the proteins tertiary structure

Answer 27

its global 3D structure (the folded structure of the entire polypeptide chain)

Question 28

the tertiary structure is formed by what interactions

Answer 28

by non-covalent (between side chains) and covalent interactions which hold the peptide chain in the 3D shape required for its function

Question 29

examples of non covalent interactions (4) (bonds)

Answer 29

- Ionic bonds, between side chains of basic and acidic amino acids. - Hydrogen bonds, between polarisable side chains of hydrophilic amino acids. - Hydrophobic bonds, between side chains of hydrophobic amino acids. - Van der Waals forces, which are weak electrostatic forces caused by temporary dipoles.

Question 30

examples of covalent interactions (bond)

Answer 30

Disulfide bonds (sometimes called disulfide bridges or cystines). These form between the sulfur atoms in cysteine side chains.

Question 31

why is primary structure to important for determine the tertiary structure

Answer 31

Side chains which are far apart in the primary peptide sequence can form the covalent interactions

Question 32

tertiary structure confers specific protein shape e.g.

Answer 32

- enzyme active sites - receptor binding sites

Question 33

tertiary structures can produce pockets which have a specific shape to allow entry and bonding of a protein’s substrate: what 2 names does this go by, and where?

Answer 33

→ In enzymes, this pocket is called the active site, and particular residues are present in the pocket whose side chains help to catalyse the reaction. → In receptors, this pocket is typically referred to as a binding site, which interacts with specific signalling molecules.

Question 34

soluble globular proteins are folded so that their […] are on the outside surface of the protein (and hence can form hydrogen bonds with water), while […] are buried on the inside of the protein

Answer 34

hydrophilic residues are on the outside hydrophobic residues are buried on the inside

Question 35

disulfide bonds are very important in tertiary structure, but not present in all proteins: they are

Answer 35

- covalent single bonds between 2 cysteines - can be intermolecular or intramolecular

Question 36

intermolecular vs intramolecular disulphide bonds

Answer 36

- Intramolecular. i.e. within a single polypeptide chain. Toxins from venomous animals often contain 3 or more disulfides. - Intermolecular. i.e. between different polypeptide chains. The best example of this is human insulin.

Question 37

Define secondary and tertiary protein structure. With reference to enzyme active sites, explain why tertiary protein structure is important for protein function.

Answer 37

**Define:** - Secondary – local 3D structure, backbone, hydrogen bonding - Tertiary – global 3D structure, amino acid side chains **Explain** why it’s important for enzymes: - 3D shape enables protein to perform its function - 3D structure creates active site for catalysis - Active site has shape which allows substrate access and binding - Amino acid side chains in the active site catalyse the reaction

Question 38

List 4 types of interaction which contribute to the formation of tertiary protein structure. For each interaction, suggest which type of amino acid (basic, acidic, hydrophobic, hydrophilic) usually forms these interactions.

Answer 38

**Interaction** AND **which amino acid type** - Ionic, acidic and basic AAs - Hydrophobic, hydrophobic AAs - Van der Waals, hydrophobic AAs - Hydrogen bonding, hydrophilic AAs Disulfide bond, hydrophilic AAs, exclusively Cys

Question 39

quaternary structure: not all proteins have this! - a protein has quaternary structure if it is comprised of…

Answer 39

two or more polypeptides which associate via non-covalent interactions to form a functional complex

Question 40

each polypeptide = a subunit: together, the protein =

Answer 40

oligomeric

Question 41

oligomeric proteins are formed by what?

Answer 41

the non-covalent association of two or more polypeptide units. - the individual polypeptide subunits can be the same or different. For example, fatty acid synthase has 2 identical subunits, while haemoglobin has 2 pairs of different subunits.

Question 42

bonds involved in quaternary

Answer 42

same as tertiary: the types of interactions that hold polypeptides together (in tertiary) are the same types that can hold different chains together (in quaternary).

Question 43

The sequence of a polypeptide (primary structure) results in a particular 3D shape (tertiary structure), and this is key why?

Answer 43

this shape determines the biological **function** by providing the binding site for interaction with other proteins or molecules. **sequence → shape → function**

Question 44

**protein functions → seven different categories:** - what are they?

Answer 44

- 1. **Catalysis** - enzymes which catalyse chemical reactions. 2. **Genomic** - DNA-associated proteins such as histones and transcription factors which regulate chromosome structure and gene expression. 3. **Defence** – proteins such as antibodies which are produced by the immune system to fight infection and remove foreign substances. 4. **Movement** - contractile proteins (actin and myosin) for muscle contraction, motor proteins such as kinesins to move things around within cells (molecules or whole structures such as vesicles). 5. **Structure** – proteins which define cell shape and support tissues e.g. proteins such as collagen and elastin in connective tissues. 6. **Transport** – proteins which carry molecules around the body (e.g. haemoglobin), and membrane proteins which control transport of molecules and ions in and out of cells. 7. **Signalling** – hormones which convey signals between cells, and receptors which receive the signals and trigger signalling pathways (themselves involving chains of interacting proteins).

Question 45

Enzymes have substrates which bind at the

Answer 45

active site

Question 46

Receptors have ligands which bind at the

Answer 46

binding site

Question 47

Antibodies have antigens which bind at the

Answer 47

binding site

Question 48

how do enzymes function as catalysts:

Answer 48

- Biological catalysts → Lower activation barrier for chemical reaction to occur - Folding of the enzyme produces the active site where catalysis occurs → Specific shape to allow substrate access → Contains amino acids involved in substrate binding and catalysis - e.g. HMG-CoA reductase: enzymes are important drug targets

Question 49

how do they function genomically as histones:

Answer 49

- role to condense nuclear DNA into chromatin - histones = basic proteins (positively charged) which associate with negatively charged DNA and interact with it favourably → Neutralise DNA charge → Positive residues are acylated to mask their charge, reducing their interaction with the negative DNA for DNA to be revealed for transcription and replication

Question 50

how do they function as antibodies in defence?

Answer 50

- Involved in immune response, now often used for cancer treatment → Neutralise large invading molecules e.g. viruses - comprised of 4 separate proteins: have a defined quaternary structure held together by disulfide bonds - Antibodies have highly specific binding to one antigen (invading molecule) → Antigen-binding site has complementary 3D shape to the antigen Antibodies bind proteins in invading viruses or bacteria and ‘label’ them for destruction by white blood cells

Question 51

how do they function in movement as actin/myosin:

Answer 51

- Responsible for muscle motion - Myosin is a molecular motor protein → **uses energy from ATP hydrolysis to power muscle contraction**: how it does this -  energy from ATP hydrolysis to form and break bonds with actin filaments (pink), causing the myosin to slide along the actin filament. This causes muscle contraction. - head group 3D structure enables actin binding and contains a site for ATP binding and hydrolysis

Question 52

how do they help structurally as collagen

Answer 52

- Collagen has a mechanical and supportive function: its present in all connective tissues - Maintains shape of cells and tissues - Form insoluble fibres (not globular) → its grouping is called a ‘Super-secondary structure’ 1. Triple helix 2. Helices covalently crosslinked (Collagen forms a coiled coil (like a rope of 3 strands). Each strand is an alpha helix, and the 3 strands wind around each other to form the fibre. Each strand is also crosslinked to the others via lysine side chains.)

Question 53

how do they act in transport as haemoglobin

Answer 53

- Specific 3D shape to selectively bind the molecule to be transported, e.g.: 1. Cytochrome c is involved in energy generation in mitochondria and binds heme. 2. Transport proteins have specific shape to allow passage of specific compounds or ions. 3. Haemoglobin has a heme bound in every subunit, which are able to bind gases for transport.

Question 54

why is hb shape so critical?

Answer 54

Haemoglobin is a globular protein which is ordinarily soluble. Each of it’s subunits contain heme.. In sickle cell anaemia, a mutation in the β subunit causes haemoglobin to aggregate. This aggregation causes red blood cells to adopt a sickle shape, and these misshapen red blood cells block narrow blood vessels and interfere with blood flow. Hb binds and transports O2 and CO2.

Question 55

how do they act in signalling:

Answer 55

Cell signalling is a complex process which is mediated by the interaction of ligands or hormones with their specific receptors, resulting in a physiological response. **Ligands or hormones interact with the complementary 3D binding site of their receptors.** The receptor binding site contains amino acids which form specific interactions with a particular ligand or hormone. This enables binding. - **Hormones**: usually peptides (with defined secondary structures / up to ~50 residues) → Chemical messengers → e.g. insulin, glucagon, vasopressin, oxytocin - **Receptors**: often span membranes → Transmembrane region* formed of α helices with hydrophobic amino acids on the outside of the helix to interact favourable with hydrophobic lipid chains in the membrane - Ligand/hormone binding causes change in receptor shape, triggering pathway to lead to physiological responses in the cell

Question 56

what is the transmembrane region:

Answer 56

part of the receptor which traverses the membrane

Question 57

denaturation - what structure does this not disrupt?

Answer 57

doesn’t disrupt primary structure. Primary structure is only disrupted by enzymatic digestion or very extreme pH.

Question 58

what is denaturation actually then? describe it

Answer 58

the loss of tertiary and secondary protein structure caused by heat, pH, UV, heavy metals, organic solvent: → Weakening of non-covalent interactions → Breaking of hydrogen bonds

Question 59

when can denaturation be reversible

Answer 59

when mild

Question 60

when can denaturation be irreversible

Answer 60

Harsh conditions can cause irreversible denaturation: this is because: secondary and tertiary structures are broken = protein unfolds, often exposing parts of the protein which previously were buried in the interior. This can lead to the formation of new covalent bonds, and the protein will not be able to regain its native structure.

Question 61

Give ONE example of a potential cause of protein denaturation. With reference to the effect on secondary and tertiary structure, explain what happens when a protein denatures.

Answer 61

**Cause**: - Any environmental factor mentioned in lecture is acceptable e.g. heat, UV, pH, metals, organic solvent **Explain**: - Causes loss of secondary and tertiary structure - By weakening/breaking non-covalent interactions which form the secondary and tertiary structure