Proteins

Question 1

structure of protein

Answer 1

• consist of elements carbon, hydrogen, oxygen, nitrogen and sulfur (in some cases)
• specific 3d conformation
• r groups

Question 2

polypeptide definition

Answer 2

A linear sequence of
amino acids covalently joined together by peptide bonds

Question 3

difference between fibrous and globular shape protein

Answer 3

ref to page 5

Question 4

how are proteins classified

Answer 4

1. simple protein - consist of only amino acid (albumin, globulins, histones)
2. conjugated protein
   - protein combined with non-protein component, cofactor
   - cofactor aids protein function, can be inorganic or organic in nature
   - organic cofactor tightly bound to a protein-> prosthetic group
   (glycoprotein, chromoprotein, lipoprotein, flavoprotein, nucleoprotein)

Question 5

types of amino acids?

Answer 5

essential amino acids - obtained through diet
non-essential amino acids - can be synthesised by body
derivatives - DNA does not code for them, modified after incorporation into polypeptide chain
note: both are impt, essential no more impt as non essential

Question 6

structure of amino acid

Answer 6

carbon covalently attached to:
1. a basic amine group (–NH2)
2. an acidic carboxyl group (–COOH)
3. a hydrogen atom and
4. a variable group known as the R group which gives ‘uniqueness’ to the amino acid (also called the side chain)

Question 7

properties of amino acid

Answer 7

1. colourless and crystalline solids, relatively high mp
2. able to form zwitterions
3. able to act as buffer
4. unique properties of R group -> R groups have important physical and chemical properties, which influence physical and chemical properties of amino acids and protein

Question 8

How are zwitterions formed

Answer 8

• carboxyl group (–COOH) loses a hydrogen ion (H+), making it negatively charged (–COO-).

this hydrogen ion (H+) associates with the amine group (–NH2), making it positively charged (–NH3+).

resulting amino acid contains one positive charge and one negative charge, it is an electrically neutral, dipolar ion -> zwitterion

Question 9

why are amino acids insoluble in organic solvents but soluble in water? (same as carbs)

Answer 9

amine and carboxyl group of amino acids can readily ionise

Question 10

what is a buffer?

Answer 10

buffer: substance that can resist changes in pH in a solution when small amounts of an acid or alkali is added to it.

Question 11

how are amino acids able to act as buffer

Answer 11

amino acids exist as zwitterions in aq medium -> amphoteric (both acidic and basic properties)
When acid is added, an amino acid (+H3N–RCH–COO-) takes up a hydrogen ion (H+) and becomes +H3N–RCH–COOH - the carboxyl group accepts the hydrogen ion.

When alkali is added, an amino acid (+H3N–RCH–COO-) loses a hydrogen ion and becomes H2N–RCH–COOi.e. the amine group loses a hydrogen ion which combines and neutralises the OH-

amphoteric is not amphiphatic(hydrophillic hydrophobic)

Question 12

how are amino acids classified based on chemical properties

Answer 12

1. amino acid with non-polar R groups
   * R groups of amino acids are hydrocarbon in nature (C-C and C-H bonds)
   * hydrophobic and unreactive
   * localised in the interior, ie shielded from aq medium of the polypeptide as it folds into its 3D conformation
2. amino acid with polar R groups
   * polar R groups (-OH and -NH) with no net charge
   * hydrophillic
3. amino acids with charged R groups
   * have negatively charged or positively charged R group -> hydrophilic
   * Acidic amino acids have carboxyl group in R group -> net negative charge
   * basic amino acids have amine group in R group -> net positive

Question 13

what is a peptide bond

Answer 13

covalent bond formed between amine group of one amino acid and carboxyl group of the other.
process of formation of peptide bond -> condensation / dehydration rxn, water molecule eliminated

Question 14

structure of polypeptide

Answer 14

• many amino acids joined together by peptide bonds in a specific linear amino acid sequence
• Each amino acid residue forms of two peptide bonds, linked to its neighbours in a head-to-tail fashion
• Each amino acid member in a polypeptide is now known as an amino acid residue

features of polypeptide:
* a free amine group, which marks the beginning of the polypeptide – the N terminus
* a free carboxyl group, which marks the end of the polypeptide – the C terminus
* R group (side chain) of each amino acid residue projects from the backbone of the polypeptide

Question 15

property of polypeptide

Answer 15

1. presence of the free amine and carboxyl group -> ability to buffer solutions, although not to as great an extent as free amino acids
2. R groups of some amino acids can ionise -> additional buffering capacity, essential in biological systems, where small changes in pH can affect the functioning of enzymes and other proteins.
3. variations in the length and the amino acid sequence of polypeptides contribute to
   the diversity in the shape and biological functions of proteins

Question 16

biuret test (principle, method, observation)

Answer 16

principle: biuret test detects peptide bonds, all proteins (NOT amino acids) give a positive result.
Nitrogen atoms in peptide bonds complexes with Cu2+ ions to give purple colouration.

method: Add equal volume of 5% potassium hydroxide solution to test solution.
Add 1% copper sulfate solution dropwise.
Mix the contents by shaking and leave for 3 minutes (no heating required)

observation:
purple / violet colour -> presence of peptide bonds
blue colour -> which is due to the copper sulfate solution indicates the absence of peptide bonds.

Question 17

What is the primary structure of a protein?

Answer 17

the
1. unique number and
2. linear sequence of amino acids that constitute the polypeptide chain

Question 18

how does primary sequence determine protein’s structure and function?

Answer 18

• sequence of amino acids contains information necessary to specify how a polypeptide chain coils and folds into a specific 3D conformation + how each polypeptide chain will interact with another
• The sequence of the amino acids in the protein influence the characteristics of a protein (not amino acid composition)
• size charge, polarity or hydrophobicity of amino acid R groups -> determine the type and location of bonds present at higher levels of organisation in the protein -> affect the ultimate 3D conformation hence function of the protein
  e.g. sickle cell anaemia

Question 19

secondary structure

Answer 19

• hydrogen bonds at regular intervals along polypeptide backbone -> regular coiling and folding of regions in polypeptide chain -> repeated patterns
• each hydrogen bond is formed between N-H group of one amino acid and C=O group of another amino acid (doesn’t involve R groups)
• each hydrogen bond is individually weak, but collectively serve to stabilise structure
• types of secondary structure: a-helix and b-pleated sheet

Question 20

shape of a and b helix

Answer 20

shape of a helix - extended spiral spring
shape of b helix - extended zigzag, sheet-like conformation

Question 21

structure of a-helix

Answer 21

nature of bonds:
- stabilised by intrachain hydrogen bonds, which occur between C=O and N-H groups of the peptide backbone
- hydrogen bond is formed between the O atom of the C=O group of an amino acid residue (nth) and the H atom of the N-H group of another amino acid that is situated four amino acid residues (nth + 4 residue) ahead in the linear sequence.
- hydrogen bonds formed are parallel to the main axis of the helix + all C=O and N-H groups of the peptide backbone can participate in hydrogen bonding -> maximum stability to the α-helix.

R groups:
- R groups of the amino acid residues project outside the helix, perpendicular to the main axis -> prevent steric interference with the polypeptide backbone and with each other.
- The chemical property of R groups (hydrophilic or hydrophobic) will influence the way the α helix interacts with the surrounding medium or other proteins
- Proline and hydroxyproline insert a kink and disrupt the formation of the α-helix.
- Amino acids with bulky R groups like tryptophan, if present in large numbers can also interfere with the formation of the α-helix

Question 22

b pleated sheet structure

Answer 22

nature of bonds
- also stabilised by hydrogen bonds (occur between C=O and N-H groups of polypeptide backbone)
- hydrogen bonds can occur between C=O and N-H groups withing same polypeptide chain / neighbouring polypeptide chain

r groups:
- usually small R groups as bulky R groups interfere with formation of b-pleated sheet by causing steric hindrance

b pleated sheet comes in 2 varieties
- antiparallel b-pleated sheet -> neighbouring hydrogen-bonded polypeptide segments run in opposite N-terminus to C-terminus directions
- parallel b-pleated sheet -> hydrogen bonded segment sun in the same N terminus to C terminus direction

Question 23

where can hydrogen bonding occur in b pleated sheet

Answer 23

1. occur among two or more segments of same polypeptide chain - intrachain sheet
2. occur among two or more segments of different polypeptide chains - interchain sheet

Question 24

What is a protein

Answer 24

Molecules made up of one or more polypeptide chains, constructed by a set of 20 different amino acids encoded by DNA that has attained a stable, specific 3D conformation and is biologically functional

Question 25

tertiary structure definition

Answer 25

further bending, twisting and folding of polypeptide chain with secondary structures to give an overall specific 3D conformation of a protein

Question 26

tertiary structure bonds

Answer 26

- specific 3d conformation is determined by 4 types of R group interactions formed between amino acid residues some distance apart on the same polypeptide chain - R group interactions -> cause folds, bends, loops in polypeptide -> interaction between different regions of polypeptide 1. non covalent interactions (relatively weak) - ionic bonds - hydrogen bonds - hydrophobic interactions 2. covalent bonds (strong) - in particular, disulfide bonds

Question 27

hydrogen bonds

Answer 27

- formed between electropositive hydrogen atom such as that attached to nitrogen or oxygen (H atom in -OH or -NH groups) and another electronegative atoms (N or O atom) within polypeptide chain - H atoms -> partial positive charge, N and O atoms -> partial negative charge, opp charged atoms are attracted to ach other to form hydrogen bond - weak but large number of hydrogen bonds confer stability

Question 28

ionic bonds

Answer 28

- Some amino acid R groups are positively charged (e.g. NH3+ group) while others are negatively charged (e.g. COO- group) - These oppositely charged R groups may form ionic bonds - ionic bonds are relatively weak in the aqueous cellular environment and may be broken by changes in the pH of the surrounding medium.

Question 29

hydrophobic interactions

Answer 29

- polypeptide folds so as to shield hydrophobic R groups from the aqueous environment. - Interactions occur between hydrophobic R groups of amino acid residues

Question 30

disulfide bond

Answer 30

- covalent bond - formed by oxidation of sulfhydryl groups (-SH) of any two cysteine residues in the dame polypeptide chain (intrachain) or in different chains (interchain) (the -SH of each amino acid becomes S-S as H atom is lost) - breaks during reduction

Question 31

what is a domain

Answer 31

- defined as a discrete, locally folded unit of tertiary structure that usually has a specific function. - a distinct structural unit

Question 32

quaternary structure definition

Answer 32

the overall protein structure that results from the association of two or more polypeptide chains to form a functional protein.

Question 33

structure of protein? nature of bonds?

Answer 33

- each individual polypeptide adopts tertiary structure, called a protein subunit - multimeric protein -> protein with more than one polypeptide / subunit - nature of bonds: same types of interactions that produce tertiary structure also contribute to quaternary structure - include ionic bonds, hydrogen bonds, hydrophobic interactions and disulfide bonds between R groups of different protein subunits.

Question 34

hemoglobin function

Answer 34

- oxygen-binding protein found in vertebrates - found in blood within red blood cells (erythrocytes) - globular protein with a quaternary structure - function: transport oxygen (O2) in the blood from the lungs to other tissues in the body, in order to supply cells with the O2 required for aerobic respiration.

Question 35

structure of haemoglobin

Answer 35

multimeric protein comprising 4 polypeptide chains, namely 2 α-chains and 2 β-chains It is a tetramer (4 subunits, α2β2) made up of two identical dimers

Question 36

explain struture of individual subunits (primary, secondary, tertiary structure) + relate structure of protein to its function

Answer 36

primary structure: Each ab dimer is made out of two types of polypeptide chains (subunits): an α-chain (141 amino acids) and a β-chain (146 amino acids) Secondary structure: - Each polypeptide chain consists of eight α-helices (named by letters A to H, starting from the N-terminus) connected by non-helical segments - **Hydrogen bonds stabilise the eight α-helices.** Tertiary structure: Structure: Each polypeptide chain is folded such that amino acid residues located at the **surface of a subunit are generally hydrophilic while those buried in the interior of the molecule are mostly hydrophobic.** Function: haemoglobin **soluble in an aqueous medium** and hence, a good transport protein for oxygen in blood. Structure: **Folding of the polypeptide chain** also allows the formation of a **hydrophobic cleft** (lined with hydrophobic amino acid residues) to allow for the **haem prosthetic group to bind.** Each polypeptide chain also contains a haem prosthetic group. Function: Each haem group will bind 1 molecule of O2. Therefore, each haemoglobin molecule will bind to 4 molecules of O2.

Question 37

structure and function of haem prosthetic group

Answer 37

Structure: The haem group consists of an iron ion (Fe2+) held in a porphyrin ring. Relating Structure to Function: The iron ion, Fe2+, binds one of the oxygen atoms in a molecule of O2. The Fe2+ can combine reversibly with O2 -> enhances the release of O2 in metabolically active tissues such as muscles.

Question 38

structure and function of haemoglobin molecule (quaternary structure)

Answer 38

Quaternary structure - The two subunits in each dimer are held together primarily by hydrophobic interactions, but ionic and hydrogen bonds also occur. - the four subunits form a globular molecule that is held by multiple non- covalent interactions. function: - Fe2+ in the first haemoglobin subunit binds 1 molecule of O2, the F helix is pulled closer to the haem group. - This pull creates a strain on the other haemoglobin subunits, such that the previously obscured haem groups of the other subunits are revealed -> remaining subunits changed their 3D conformation slightly -> affinity for O2 molecules increases, allowing their respective haem groups to bind O2 more readily, - Therefore, haemoglobin is known as an allosteric protein, and this mechanism of oxygen binding is known as cooperativity / cooperative binding ## Footnote extra info: allosteric protein is one in which the binding of a ligand, e.g. O2 in the case of haemoglobin, to onesite affects the binding properties of another site on the same protein.

Question 39

oxygen dissociatin curve of haemoglobin and myoglobin

Answer 39

haemoglobin: - sigmoidal shape of curve as haemoglobin is made up of 4 polypeptides, which displays subunit cooperativity - oxygen loaded haemoglobin in the lungs where the partial pressure is high, and unloaded from haemoglobin in the rest of the body tissues where the partial pressure is low, - when PO2 (partial pressure O2) is high, more haemoglobin subunits are bound to O2 and the haemoglobin is more saturated with O2. The reverse occurs when PO2 is low. myoglobin (stores O2 in muscle cells) - Myoglobin is made up of only one polypeptide -> oxygen-dissociation curve of myoglobin is hyperbolic. - Myoglobin has a higher affinity binding to oxygen (i.e. more saturated with oxygen) than haemoglobin at the same partial pressure. - Subunit cooperativity thus allows haemoglobin to be an efficient oxygen carrier, since haemoglobin loses oxygen quickly in an environment with low oxygen concentration (eg muscles) and vice versa

Question 40

function of collagen

Answer 40

- collagen's strong, insoluble fibres are the major stress-bearing components of connective tissues - great tensile strength -> provide bones with structure, flexibility and strength, tendons with strength and structure required to transmit muscular contraction and veins, arteries and capillaries the strength, structure and flexibility to withstand blood pressure.

Question 41

structure of collagen molecule

Answer 41

primary structure: - A single collagen molecule consists of three polypeptide chains. - The amino acid sequence of a collagen polypeptide consists of a repeating tripeptide sequence of Glycine – X – Y where (i) X is often proline (prevent it from forming a-helix) (ii) Y is often hydroxyproline or hydroxylysine - Each chain is about 1000 amino acid residues long. secondary structure: - Each collagen polypeptide assumes a left-handed helical conformation with about three residues per turn known as the collagen helix - This regular repeated structure - secondary structure, indicative of a fibrous protein - Each of these polypeptide chains is called an α-chain. Quaternary structure: - Three parallel α-chains wind around each other with a gentle, right-handed, rope-like twist (i.e. right-handed triple-helix) to form a tropocollagen - well-packed, rigid triple-helix structure is responsible for its tensile strength - twist in helix cannot be pulled out under tension as its component polypeptide chains are supertwisted about each other

Question 42

features of tropocollagen

Answer 42

- Every third residue of each polypeptide passes through the centre of the triple-helix, which is so crowded that only the small R group of glycine (Gly) can fit in -> explains absolute requirement for Gly at every third residue -> allows the three helical α-chains to pack tightly together, providing high tensile strength. - the residues in the X and Y positions are located on the outside of the triple-helix, where there is room for the bulky R groups of proline and other residues. - proline with its ring structure, stabilises the rigid three-stranded collagen helix. - covalent cross-links are also present within tropocollagen molecules to further impart the collagen fibre with high tensile strength. tropocollagen is held together by an extensive network of hydrogen bonds. - Hydrogen bonds formed between the N-H group of Gly residue in one a-chain and the C=O group of another amino acid residue in a neighbouring a-chain help hold the three chains together. - The hydroxyl groups (–OH) of hydroxyproline and hydroxylysine residues also participate in interchain hydrogen bonding.

Question 43

formation of a collagen fibre

Answer 43

- Many tropocollagen molecules lie side by side, linked to each other by covalent cross-links between the carboxyl end of one molecule and the amino end of another -> gives rise to a structure known as a collagen fibril - accumulated covalent cross-links in collagen fibrils -> results in increasingly rigid and brittle character of aging connective tissue - tropocollagen molecules are arranged in a staggered manner with each other, arrangement is stabilised by hydrophobic interactions between tropocollagen molecules -> confers collagen greater strength. - aggregation of collagen fibrils form a collagen fibre

Question 44

bonds located in haemoglobin and collagen

Answer 44

haemoglobin - mainly hydrophobic interactions, but ionic bonds and hydrogen bonds do occur collagen - hydrogen bonds (between N-H group of Gly residue in 1 chain and C=O group of another amino acid residue in neighbouring chain), covalent cross links(between carboxyl end of one molecule and amino end of another), hydrophobic interactions (btw tropocollagen molecules)

Question 45

denaturation

Answer 45

definition: loss of specific 3D conformation of a protein molecule - happens when bonds that maintain conformation is broken, protein unfolds, can no longer perform biologicla function - temp or permanent (most cases) - primary structure (amino acid seq) is unaffected

Question 46

what is renaturation

Answer 46

when denatured globular proteins regain specific 3D confomation and biological activity if returned to conditions in which 3D conformation is stable (but rare)

Question 47

what does denaturation disrupt

Answer 47

disrupt secondary, tertiary and quarternary structures disrupts 1. R group interactions (disulfide bonds, ionic bonds, hydrogen bonds, hydrophobic interactions) 2. hydrogen bonds formed between N-H and C=O groups of polypeptide backbone (rmb secondary structure)

Question 48

denaturants and their effects

Answer 48

1. Heat - Excessive heat increases vibrations of the atoms, leading to disruption of hydrogen bonds, ionic bonds and hydrophobic interactions. Application: egg white becomes opaque during heating as denatured proteins are insoluble and solidify. 2. Changes in pH - Drastic changes in pH changes the charges in the acidic and basic R groups, leading to disruption of ionic bonds and hydrogen bonds. 3. Organic solvents - Transfer of a protein from an aqueous environment to an organic solvent can disrupt hydrophobic interactions that make up the stable core of globular proteins. The protein turns inside out and the hydrophobic regions changes place with the hydrophilic regions. 4. Urea / Detergents - Addition of chemicals can disrupt ionic and hydrogen bonds that maintains the protein’s conformation.