BG7004: Advanced Cell Biology

Lecture 2: Protein structure and function

Learning Outcomes:

• Shape and structure of proteins

• How proteins work

• How the functions of proteins are controlled

Dr Noreen Ishak

Amino acids are the subunits of proteins or

polypeptides

There are 20 amino acids which are grouped into 4 classes

according to their side chains

hydrophilic hydrophobic

Amino acids are joined together by a peptide bond

Condensation

Proteins are built up by amino acids that are linked by peptide bonds to form a polypeptide chain

The shape of a protein is determined by its amino acid sequence

Each type of protein differs in its sequence and number of amino acids hence the sequence of the chemically different side chains make each protein different from each other

3D structure of a protein is stabilised by noncovalent bonds

between different parts of the protein

Ionic bonds: Formed when electrons are transferred from one atom to the other

Hydrogen bonds: Formed by electrical attractions between a positively charged hydrogen atom in one molecule with a negatively charged oxygen or nitrogen in another molecule

Van der Waals attractions: Formed by electrical attractions between two atoms that are close to each other

Hydrophobic interaction: Hydrophobic groups are forced together by repulsion from water

Four types of noncovalent bonds important in

biological systems

Three types of noncovalent bonds help proteins to fold

Hydrophobic forces help proteins fold into compact

conformations

Polar amino acid side chains tend to gather on the outside of the folded protein where they can interact with water

Nonpolar amino acid side chains are buried inside to form a highly packed hydrophobic core of atoms that are hidden from water

Most proteins are folded by noncovalent bonds into only

one stable conformation in which free energy is minimised

Protein can be denatured and renatured

When protein folds improperly, they can form aggregates

that can damage cells even whole tissues

Aggregated proteins have been associated with diseases such as Huntington’s disease, Alzheimer’s disease and prion disease

In the case of prion disease, prion protein can adopt a special misfolded form that is considered ‘infectious’ due to its ability to convert normal protein into an abnormal conformation

Proteins are in different shapes and sizes

Globular shape Fibrous shape

Primary structure (amino acid sequence)

Secondary structure (α helices, β sheets)

Tertiary structure (3D structure of a polypeptide)

Quartenary structure (3D structure of a protein complex consisting of more than one polypeptide)

Four levels of organization in the structure of a

protein

α helix – generated when a single polypeptide chain

turns around itself to form a cylindrical structure

In an α helix, a hydrogen bond is made between every fourth

peptide, linking the C=O of one peptide bond to the N-H of

another

Video Clip: 04.2 Alpha helix

Hydrogen bond

Peptide bonds Peptide bonds

Lipid bilayer

oxygen

Video Clip: 04.4 Beta sheet

The β sheets are made when hydrogen bonds form

between segments of polypeptide chains lying side by

side

Both types of β sheets are common in proteins

Antiparallel β sheet

Parallel β sheet

Many proteins are composed of separate functional domains made with α helix and β sheet

Proteins can bind to each other through the binding sites

on the surface of a protein to form dimers or multimers

Proteins can assemble into filaments, sheets or spheres

based on the differences in their binding sites

Extracellular proteins such as collagen are stabilised by

cross linking through disulfide bonds

Collagen is a triple helix formed by three extended protein chains that wrap around one another

Many rod-like collagen molecules are cross-linked together in the extracellular space to form collagen fibrils that have the tensile strength of steel

Video Clip: 04.6 Disulfide bonds

Extracellular proteins such as collagen are stabilised by cross linking through disulfide bonds

Disulfide bonds can form between adjacent cysteine side chains

A disulfide bond can have a major stabilising effect on a protein since it requires high energy to be disrupted

General protein functionsProteins have many functions such as:

Enzyme – catalyses enzymatic reactions (pepsin, DNA polymerase)

Structural protein – provides mechanical support (collagen, elastin)

Transport protein – carries small molecules or ions (haemoglobin, transferrin)

Motor protein – generates movement in cells (myosin, kinesin)

Storage protein – stores small molecules or ions (ferritin, casein)

Signal protein – carries signals from cell to cell (insulin, epidermal growth factor)

Receptor protein – detects signals and transmits them to the cell’s response machinery (acetylcholine receptor, insulin receptor)

Gene regulatory protein binds to DNA to switch genes on and off (lactose repressor)

All proteins must bind to particular ligands to

carry out their various functions

Any substance that can bind to a protein is referred to as a ligand for that protein

A ligand can be an ion, a small molecule, or a macromolecule

The binding between a protein and its ligand is highly selective and requires the formation of noncovalent bonds

Example 1 - Antibody

Video Clip: 04.7 Antibodies

Example 1 – Antibody can bind tightly with its

antigen through the binding site

Example 2 – Enzymes bind to substrates and convert

them into chemically modified products

Lysozyme cleaves a polysaccharide chain

Video Clip: 04.8 Lysozyme reaction

1) The catalytic activity of enzymes in cells are often regulated by other molecules in several ways:

The production of an enzyme can be regulated by gene expression

The availability of an enzyme can be controlled by subcellular localization of the protein

The activity of an enzyme can be negatively regulated by feedback inhibition

The enzyme activity can be stimulated by a regulatory molecule through a positive regulation

Mechanisms that regulate the activity of

proteins and enzymes

In this negative feedback inhibition, the end product Z inhibits the production of the first enzyme that is required for the synthesis of X and thereby controls the concentration of its final product

Feedback inhibition can work almost instantaneously and is rapidly reversedwhen product levels fall

Feedback inhibition regulates the flow

through biosynthetic pathways

Many enzymes have two binding sites: one for the

substrate and the other for the regulator (negative

control)

Aspartate transcarboxylase is an allosteric enzyme from E.coli. It catalyses an important reaction that begins the synthesis of the pyrimidine ring of C,U and T nucleotides. One of the final products of this pathway, cytosine triphosphate (CTP), binds to the enzyme to turn it off whenever CTP is plentiful. CTP can thus act as a negative regulator. The binding of CTP changes the protein confirmation and inactivates the enzyme.

In a positive regulation, a regulator (ADP) can bind

to the enzyme to lock them in the active form

2) Protein phosphorylation is a very common mean to

regulate protein activity

Thousands of proteins in a typical eukaryotic cell are modified by the covalent addition of a phosphate group

A phosphate group is transferred from ATP to an amino acid side chain of the target protein by a protein kinase

Removal of the phosphate group is catalyzed by a protein phosphatase

The phosphorylation of a protein by a protein kinase can either increase (ON) or decrease (OFF) the protein’s activity depending on the site of phosphorylation and the protein structure

3) The activity of GTP-binding protein is also regulated by

the cyclic gain and loss of a phosphate group

GTP-binding proteins can function as molecular switches

The activation of a GTP-binding protein generally requires the presence of a tightly bound GTP molecule (switch on)

Hydrolysis of this GTP molecule by GTPase produces GDP and inorganic phosphate (Pi), and it causes the protein to convert to an inactive conformation (switch off)

Resetting the switch requires the tightly bound GDP to dissociate, a slow step that is greatly accelerated by specific signals

Once GDP dissociates, a molecule of GTP quickly replaces it and returns the protein to its active conformation

Guanosine Triphosphate (GTP)

4) ATP hydrolysis allows motor proteins to produce large

movement in cells

ATP binding shifts a motor protein from confirmation 1 to 2

The bound ATP is then hydrolysed to produce ADP and inorganic phosphate (Pi), causing a change from confirmation 2 to 3

The release of the bound ADP and Pi drives the protein back to confirmation 1

ATPase

Summary

Each type of protein has a unique amino acid sequence which determines both its

3D shape and biological activity

The folded structure of a protein is stabilized by multiple noncovalent interactions

between different parts of the polypeptide chain

There are four levels of organisation in the structure of a protein

Activities of most enzymes are strictly regulated

Properties Fibrous protein Globular protein

Shape Long and narrow Rounded/spherical

Role Structural (strength and support) Functional (catalytic, transport, etc)

Solubility (Generally) insoluble in water (Generally (soluble in water)

Sequence Repetitive amino acid sequence Irregular amino acid sequence

Stability Less sensitive to changes in heat, pH, etc More sensitive to changes in heat, pH, etc

Examples Collagen, myosin, fibril, elastin, keratin, actin Catalase, haemoglobin, insulin, immunoglobulin

Chapter 4 of Essential Cell Biology

Essential concepts and Key terms

Attempt questions 10, 11, 15, 19

Follow-up tasks for Lecture 2