L12: Protein function and enzymology

What is protein structure crucial for?

Specific protein functioning

What are common protein structures that relate to their function

• Membrane proteins- receptors, transporters

• Fibrous proteins

What are enzymes absolutely specific for

Their substrate

How much do enzymes speed up reactions and how do they do it

• By factors u to 10^12

• By reducing activation energy

How many reactions do each enzyme catalyse

Enzymes can catalyse several hundreds per second

What is the active site

The part of the enzyme which they bind their substrates to and catalyse the reaction

What is induced fit

When the substrate binds to the enzyme, the enzyme undergoes a conformational change allowing it to better fit the substrate. E.g. hexokinase binding to glucose

Describe the process of substrates binding to enzymes

1. Substrates enter the active site, and the enzyme changes shape so its active site enfolds the substrate (induced fit)

2. The substrates are held in the active site by weak interactions such as hydrogen bonds and ionic bonds

3. The active site can lower Ea and speed up a reaction by acting as a template for substrate orientation, stressing the substrate and and stabilizing transition state, providing a favourable microenvironment or anticipating directly in the catalytic reaction

4. Substrate converted into products

5. Products released

6. The active site becomes available for 2 new substrate molecules

What enzymes are used in biological detergents

Proteases and lipases

Which enzyme produces high fructose corn syrup

Glucose isomerase

How can enzymes be regulated

• Proteases in the blood

• End product inhibition

• Binding of another molecule such as a phosphate

What did Frances Arnold do?

Worked out how to generate and select enzyme variants with desirable proteins eg. For uses in the chemical industry

What is the structure of antibodies

2 heavy chains and 2 light chains joined by disulfide bridges

What regions do the heavy and light chains have?

Variable region (V) and constant region (C)

What do the variable regions allow on antibodies?

Allows variation within that area of the protein so that it has the ability to bind to lots of different antigens on different epitopes

What is antigen specificity conferred by?

The variable regions

What is the epitope

The small part of the antigen in which an antibody recognises

Where does the mutation for sickle cell disease occur?

The 6th amino acid from the n terminus on the surface of the beta subunit

What happens to the beta subunit after the sickle cell mutation

It crystalises into a fibre so the red blood cell deforms and the capacity to carry oxygen is reduced

What is crucial for homeostasis

That protein function is regulated

How many subunits does haemoglobin have

4 that all interact together to form a functional protein

myoglobin

A single subunit protein that binds to oxygen with a high affinity for oxygen at a low concentration

Why does myoglobin not release oxygen as well as haemoglobin

As myoglobin has a high affinity for oxygen so the oxygen does not get released into tissues

Where is haemoglobin fully saturated

In the lungs where oxygen partial pressure is high and oxygen is fully saturating the haemoglobin

Where in the body is high and low oxygen partial pressure

• High in the lungs

• Lower in active muscles/respiring tissues

What happens to myoglobin when oxygen binds

The structure changes allowing the next oxygen to bind more easily and the structure changes again when the oxygen is released

How is haemoglobin able to release more energy

In respiring tissues, protons are released so the pH is reduced and this lowers its affinity to oxygen so the haemoglobin to release more oxygen

What can also reduce haemoglobin's affinity for oxygen

deoxyhaemoglobin can bind to BPG which will reduce

How is protein function regulated

As protein conformation is flexible, regulatory proteins can occur in 2 or more conformations with different proteins

What are the 2 structural forms of haemoglobin?

• The T state where oxygen binds poorly

• The R state where oxygen binds well

What does the transition of haemoglobin between the T state and R state allow?

It allows the binding & release of oxygen when necessary (which allows the shape of the sygmodial curve)

What increases BPG concentration

High altitude training which increases BPG concentration and increases release of oxygen from haemoglobin

Features of hexokinase

• Has 2 domains ( catalytic and regulatory)

• Activity is inhibited by glucose-6-phosphate

• It is expressed in muscles

• Controls glycolysis rate and has a high affinity for glucose

Features of glucokinase

• Has one domain

• Expressed in the liver

• Not inhibited by glucose-6 - phosphate

• Has a low affinity for glucose so requires a high concentration of glucose to work

What are the membrane spanning parts of proteins

Alpha helices (around 7-12 of them) or beta-barrels

What part of membrane proteins are hydrophobic and hydrophilic

• Hydrophobic - part exposed to membrane

• Hydrophilic - part exposed to inside and outside of cell

Functions of membrane proteins

• Cell cell recognition

• Signal transduction via hormone receptors

• Transport

What identifies our blood type

The carbohydrate group which attach to our Red blood cells

What is signal transduction

When cells (such as agonists) cannot pass through the membrane due to size or properties so they bund to the surface of a receptor protein which causes a conformational change and the signal is transduced along the membrane