Lecture01: Amino Acids and Proteins, Protein Structure and Function, Oxygen binding proteins

Are protein polymers linear or branched?

Linear, chain of amino acids

What are protein monomers?

Amino acids

What are protein polymers?

Polypeptides

How many polypeptide chains do simple proteins have?

Just one

With proteins with a prosthetic group, what is the polypeptide portion called?

Apoprotein

What does an amino acid consist of?

Central carbon
NH2
COOH
Hydrogen
R group

What is the pk value of COOH?

around 2

What is the pk value of NH2?

around 10

If the pH value is below the pk value of COOH, do you have a cation or anion?

Cation because the NH2 would be protonated (NH3+) as well as the COOH (acidic conditions donate the proton)

If the pH value is above the pk value of NH2, do you have a cation or anion?

Anion because COOH would be deprotonated (COO-) as well as NH2 (basic conditions take protons from the functional groups, thus deprotonating the COOH group and giving it a negative charge)

If the pH value is between the pk value of the COOH group and the NH2 group, what do you have?

A zwitterion (charge is 0)

Do L-amino acids or D-amino acids exist in nature?

L-amino acids ONLY

What is the isoelectric point?

It is the pH value at which the number of positive charges is equal to the number of negative charges for the entire protein molecule

What covalent bond connects amino acids?

A peptide bond (C-N)

What are the 2 traits of a peptide bond?

-non-ionizable
-can hydrogen bond

What forms the primary structure of proteins?

-primary structure is just the linear sequence of amino acids, so the only important bonds are the covalent peptide bonds as well as disulfide bonds

What are disulfide bonds?

-oxidation between 2 cysteine residues

Where does disulfide bond formation occur?

Endoplasmic reticulum

Which types of proteins usually have disulfide bonds?

-secreted proteins because the formation happens at the ER, thus cytoplasmic proteins usually lack disulfide bonds

What does the "real" structure of a peptide bond look like?

It is a resonance hybrid somewhere in between the two resonance structures

Can the peptide bond rotate?

no because amide has the resonance giving a double bond structure

What type of noncovalent interactions occur in the secondary structure or proteins?

Hydrogen bonds

What are the two main secondary structures that can form?

-alpha helices
-beta pleated sheets

Describe alpha helices

right handed corkscrew
-very compact
-maintained by hydrogen bonds parallel with helix axis

Which two amino acids cant be apart of alpha helices?

-proline (too rigid)
-glycine (too flexible)

Are beta sheets parallel or antiparallel?

They can be both

What is the main noncovalent interaction that is present in tertiary structures?

hydrophobic interactions within same polypeptide chain

Describe quaternary structure?

interactions between different polypeptide chains
-subunits held together by noncovalent interactions or disulfide bonds

What are proteins that assist in protein folding?

Chaperones

How are amino acids assembled?

In the N to C direction

Which amino acids can post-translationally be modified with a phosphate residue?

-serine
-threonine

(the -OH amino acids)

Which amino acids can post-translationally be modified with N-acetylgalactosamine?

-serine
-threonine

"O-linked"

What other amino acid can post-translationally be modified with N-acetylglucosamine?

-asparagine

"N-linked"

What does the solubility of proteins rely on?

-pH
-salt concentration

How does increasing salt concentration effect protein solubility?

-it increases it by neutralizing the charges on the proteins, which reduces attraction between neighboring protein molecules

What happens if the salt concentration is too high?

The proteins will precipitate because the water will become tied up with the salt molecules instead of the protein

When is protein solubility minimal in terms of pH and why?

When pH=pI because this is when the salt bonds form, resulting in protein molecules binding together into insoluble aggregates

What wavelength do peptide bonds absorb light?

190 nm

What wavelength do amino acids absorb light because of aromatic side chains?

280 nm

What separates proteins by size?

Dialysis

What separates proteins by charge and how?

Electrophoresis, separates by charge-mass ratio

-negatively charged proteins move to anode
-positively charged proteins move to cathode

What are the 2 main shapes of proteins?

1) globular - compact, water soluble, hydrophobic core
2) fibrous - long, thread-like, structural functions (keratin)

Ways to destroy higher order structures of proteins?

-cleavage of peptide bonds
-oxidizing/reducing agent to get rid of disulfide bonds
-get rid of noncovalent interactions through heating, detergents, acids/bases, heavy metal ions

Does denaturation change physical properties?

Yes

Is denaturation reversible or irreversible?

irreversible

What do denatured proteins refold into?

Fibrils

What does refolding often lead to?

misfolding

What are amyloids?

aggregation of misfolded proteins (short stretches of B-pleated sheets that run perpendicular to the axis of the fibril)

In Parkinson's disease, what happens?

The misfolded alpha-synuclein protein aggregates into lewy bodies

what do senile plaques contain in alzheimer's patients contain?

aggregates of B-amyloid, which eventually leads to hyperphosphorylation of the tau protein

How can protein misfolding be contagious?

If "scrapie" product of protein folding forms, then other proteins can misfold as well into contagious misfolded proteins (prions)

What disease is transmittable in humans and results from prions?

Variant CJD (vCJD) - spongiform encephalopathy

What is the oxygen binding protein in blood?

hemoglobin

What is the oxygen binding protein in muscle?

myoglobin

Does myoglobin contain disulfide bonds?

No

How many subunits does hemoglobin have?

4 subunits (all of which are similar to the structure of myoglobin)

How many heme groups are in hemoglobin?

4

Which form of iron is found in heme?

Fe+2 (ferrous)

What is heme?

iron containing porphyrin ring

What binds the oxygen?

Iron

Which binds oxygen more tightly, hemoglobin or myoglobin?

myoglobin (muscles are more active and require more oxygen, thus they bind O2 tighter)

What is the curve shape of myoglobin?

hyperbolic

What is the curve shape of hemoglobin?

sigmoidal

What conformation is deoxyhemoglobin in?

The T state (tense)

What conformation is oxyhemoglobin in?

The R state (relaxed)

What allows the hemoglobin to go from the T state to the R state?

After O2 binds, the bond distance between the iron and the surrounding nitrogens shortens, this destabilizes the interaction with the other subunits, whole molecule shifts to R conformation (positive cooperativity)

Which molecule stabilizes the T state and promotes oxygen release?

2,3 BPG (negative allosteric effector)

When does 2,3 BPG concentration increase?

hypoxic conditions (like muscles), to promote release of O2 into these hypoxic tissues that need it

What is the Bohr effect?

Low pH (Increased H+) decreases hemoglobins binding affinity towards oxygen

Reasoning:

Oxygenated hemoglobin is more acidic than deoxygenated hemoglobin

Hemoglobin + O2 ----> hemoglobin-O2 + H+

With low pH (increased H+), equilibrium shifts to the left, promoting dissociation of oxygen from hemoglobin (oxygen release)

CO2 effect?

lCO2 (O=C=O) also decreases oxygen
affinity by binding covalently to amino
groups forming carbamino hemoglobin, reversible

Which has a higher binding affinity to hemoglobin, CO or O2?

CO

How is CO2 transported in the blood?

CO2 + H20 <----> H2CO3 (carbonic acid) <----> HCO3- (bicarbonate)

Enzyme to make carbonic acid?

Carbonic anhydrase

In tissues, how does CO2 get moved?

-it gets brought in

CO2 + H2O ----> H2CO3 -----> HCO3- + H+

-The H+ reduces hemoglobins binding affinity to oxygen, causing O2 to release into the tissues and bicarbonate exits the cell

In the lung capillaries, how does CO2 get moved?

-it gets exhaled out

HCO3- + H+ ----> H2CO3 ----> H2O + CO2 (released from lungs)

-hemoglobin gets saturated with oxygen at lungs