Biochemistry Ch 5 Globular Proteins

Proteins function by interacting how with other molecules?

Dynamically

What are the two types of interaction proteins can broadly have with other molecules?

• Protein acting as a reaction catalyst, or enzyme, alters the chemical configuration or composition of a bound molecule

• Neither the chemical configuration nor the composition of the bound molecule is changed

The function of many proteins involve what type of binding to other molecules?

Reversible

A molecule bound reversible by a protein is called a what?

Ligand

A ligand can be what type of molecule?

Any including another protein

What term describes the nature of protein-ligand interaction which makes them critical to life, allowing the organism to respond rapidly and reversible to changing environmental and metabolic circumstance?

Transient

A ligand binds a protein at a binding site that is complementary to the ligand in what aspects?

• Size

• Shape

• Charge

• Hydrophobic and hydrophilic character

A given protein may have one or multiple binding sites for ligands?

Multiple

Molecular interactions with proteins are specific or non-specific?

Specific

A protein can bind to one ligand or multiple?

Multiple

Proteins are flexible or rigid?

Flexible

Through what process do proteins maintain flexibility?

Conformational changes

A subtle conformational change physically affects a protein how?

Reflects molecular vibrations and small movements of amino acids residues throughout the protein

A dramatic conformational change physically affects a protein how?

Major segments move up to several nanometers

Specific conformational changes are frequently essential or non-essential for protein function?

Essential

Binding of a protein and a ligand is often coupled with what type of change in a protein that makes the binding sites more complementary to the ligand, permitting tighter binding?

Conformational change

The structural adaptation that occurs between protein and ligand is called what?

Induced fit

In a multi-subunit protein a conformational change in one subunit often does or does not affect the conformation of other subunits?

Often does affect conformation of other subunits

What is induced fit?

When ligand binding is coupled to conformational changes, sometimes quite dramatically

What is cooperativity in proteins?

In multi-subunit proteins, conformational changes in one subunit can affect the others

Is reversible binding of ligands an essential process?

Yes

Is specificity of ligands and binding sites an essential process?

Yes

Do interactions between ligands and binding sites have the ability to be regulated?

Yes

The importance of, reversible binding of ligands, is illustrated well by this globular protein that transports oxygen

Hemoglobin

What are the five main functions of globular proteins?

• Storage of ions and molecules

• Transport of ions and molecules

• Defense against pathogens

• Muscle contraction

• Biological catalysis

What are two globular proteins whose main function is storage

Myoglobin and ferritin

What are two globular proteins whose main function is transport?

Hemoglobin and serotonin transporter

What are two globular proteins whose main function is defense against pathogens?

Antibodies and cytokines

What are two globular proteins whose main function is muscle contraction?

Actin and myosin

What are two globular proteins whose main function is biological catalysis

Chymotrypsin and lysozyme

The functions of many proteins depends on interactions with what other type of biological macromolecule?

Protein

Site where ligand binds on protein is the what?

Ligand binding site

Are prosthetic groups permanently (covalently linked) bound to a protein?

Yes

Some proteins have a bound non-protein molecule which is essential for their function. What is this called?

Prosthetic group

A globular protein that transports oxygen has a prosthetic group called what?

Heme

Can small conformational changes induce a large change in conformation?

Yes, proteins are flexible

Is oxygen soluble in an aqueous solution?

Poorly soluble

Is O2 easy to diffuse over a distances greater than a few mm?

No

Oxygen is difficult to provide to the tissue in sufficient amounts why?

Poorly soluble in aqueous solution and difficult to diffuse over a significant distance

Vertebrates have evolved what two principal mechanisms for supplying their cells with a continuous and adequate flow of oxygen?

• Circulatory system

• Use of oxygen carrying proteins hemoglobin and myoglobin

What does the circulatory system do?

Actively delivers oxygen from lung/gil tissue to the rest of the organism

What is the purpose of vertebrates evolving to use hemoglobin and myoglobin?

To overcome the limitations imposed by the low solubility of oxygen in water

What are some biological problems for proteins acting as oxygen carriers?

• Protein side chains lack affinity for oxygen

• Some transitions metal bind oxygen well but would generate free radicals if free in solution which is damaging to proteins

• Fe2+ in free heme could be oxidized to Fe3+ which is very reactive

What is the biological solution to proteins side chains lacking affinity for oxygen?

Capturing the oxygen molecules using heme as protein-bound molecule

How do myoglobin and hemoglobin bind oxygen?

Protein-bound heme

What is the main function of myoglobin?

Storage of oxygen

What is the main function of hemoglobin?

Transport of oxygen

What molecules can myoglobin bind?

Oxygen and CO2

How many polypeptides is myoglobin?

One

How does myoglobin function?

Binds oxygen and CO2 from the circulatory system and transports them into the tissues. “Picks up” oxygen.

How many polypeptides is hemoglobin?

Four

How does hemoglobin function?

Carries oxygen and CO2 in the circulatory system. “Releases” oxygen.

How many binding sites for oxygen are on myoglobin?

One

What structure is myoglobin made almost entirely of?

Alpha helices

What are the two other notations for the proximal His? And what information do they tell you?

His93 - His is the 93rd residue from the amino terminal end

HisF8 - His is the eighth residue in alpha helix F

What are the two other notations for the distal His? And what information do they tell you?

His64 - His is the 64th residue from the amino terminal end

HisE7 - His is the 7th residue in alpha helix E

The proximal His is closer or further from protein bound Heme?

Closer

The distal His is closer or further from protein bound Heme?

Further

What types of polypeptide chains make up Hemoglobin?

Two alpha chains and two beta chains

In addition to being a globular protein hemoglobin is also what type of protein?

An allosteric protein

What is an allosteric protein?

One in which the activity of a binding site is affected by a different site or sites in the protein.

The affinity of hemoglobin to oxygen depends on what factors?

• Concentration of O2

• BPG

• pH

What level of structure does myoglobin have?

Tertiary

What level of structure does hemoglobin have?

Quaternary

How do the primary and tertiary structures of myoglobin and hemoglobin subunits compare?

• Low sequence similarity

• High structural similarity

What type of residues surround heme?

Hydrophobic

Iron deficiency affects the amount of oxygen we can carry how?

It lowers the amount of oxygen we can carry

Is heme a protein?

No, heme is a non-protein part of a protein known as a prosthetic group

What is the structure of heme?

A protoporphyrin IX ring with a bound iron atom in its ferrous (Fe2+) state.

Protoporphyrin IX is an example of what class of molecule?

Porphyrin

Describe the structure of protoporphyrin XI

Example of porphyrin, consists of four pyrrole rings linked by methene bridges, with substitutions at one or more positions denoted X.

The iron atom of heme has how many bonds? What type of bonds? What are these bonds attached to? Geometrically how do the bonds lay?

The iron atom of heme has 6 coordination bonds

4 bonds are attached to the porphyrin ring system and are planar relative to the ring system

1 bond is attached to the proximal His residue which is perpendicular to the ring system

1 bond is attached to the an O2 atom which is also perpendicular to the ring system

Iron in its ferric or ferrous form can bind oxygen?

Ferrous

Simultaneous reaction of an O2 can result in what conversion inside heme? But what prevents this?

Simultaneous reaction of an O2 molecule can result in the conversion of Fe2+ to Fe3+, this is prevented by proteins.

How do CO and O2 relate in shape and size?

They are similar

Between CO and O2 which molecule binds better to free heme and how much better?

CO binds 20,000x better than O2 to free heme

What does CO bind much better than O2 to free heme?

Carbon in CO has a filled lone electron pair that can be donated to vacant d-orbitals on the Fe2+

Between CO and O2 which molecule binds better to bound heme and how much better?

CO binds 250x better than O2 to bound heme

Why is CO toxic?

It competes with oxygen. It blocks the function of myoglobin, hemoglobin, and mitochondrial cytochromes that are involved in oxidative phosphorylation.

What effect weakens binding of CO to myoglobin?

Linear perpendicular arrangement is sterically blocked by His E7 (Distal His). O2 adopts a slight angle to avoid this problem.

Heme group is a strong chromophore and absorbs light from which spectrums?

Ultraviolet and visible range

Ferrous form (Fe2+) without oxygen has an intense bands at what length and what color is it?

Intense sorbet (reddish tinge) band at 429nm

When oxygen binds to ferrous (Fe2+) its intense absorption band is shifted to what length?

414nm

Binding of oxygen can be monitored by what type of spectrophotometry?

UV-Vis

Deoxyhemoglobin is found where in the body primarily and what color does it appear?

Found in venous blood appears purplish

Oxyhemoglobin is found where in the body primarily and what color does it appear?

Found in arterial blood and appears red

Ligands binds via what type of interactions?

Weak interactions such as hydrogen bonding, hydrophobic effects, and ionic interactions

ka describes what aspect of protein ligand binding?

Association rate constant

kd describes what aspect of protein ligand binding?

Dissociation rate constant

The protein ligand process will reach equilibrium at what point?

When ka=kd (association and dissociation rates are equal)

ka[P]*[L]=kd[PL]

Ka describes what aspect of protein ligand equilibrium?

Equilibrium association constant

Kd describes what aspect of protein ligand equilibrium?

Equilibrium dissociation constant

How can Ka be calculated?

Ka = [PL]/([P]*[L]) = 1/Kd

(Product / Reactant)

What is the relationship between Ka and Kd

Inverse

Does a protein ligand reaction ever reach complete binding meaning all proteins are bound by ligand?

No

Fraction of occupied binding sites is denoted by what two symbols?

Y or theta

How can Y (fraction of occupied binding sites) be calculated?

Y=[L]/([L]+Kd)

The fraction of bound sites on a protein depends on what 2 factors?

Free ligand concentration and Kd

What is Kd’s relationship to [L]?

Kd is the [L] at which half of the ligand-biding sites are occupied

When a ligand is a gas binding is expressed in terms of what?

Partial pressures