BIOC192 - module 1 and 2

Haemoglobin in red blood cells in the blood n eeds to be able to :

-Binding O2 in the vicinity of the lungs where pO2 is ~100 torr
-Release the O2 in the vicinity of peripheral tissues where the pO2 is ~20 Torr.
Haemoglobin is therefore evolved to bind O2 LESS tightly than myoglobin. Can change shape

Protein

Non-branching polymers that form macromolecules about 50-100A. 1A=1-^-10m. A bag of lipids and nucleic acids

Each protein has specific sequence of amino acids joined together by chemical peptide bonds.
20 amino acids

e.gs: table salt, cheese,sucrose, hair, haemoglobin

Atomic structure determination of proteins

Protein crystallography
Cryo-electron microscopy
NMR spectroscopy.

Cell signaling-insulin

-Hormone insulin.
Generated and binds insulin receptor to signal cell to take up glucose.
Structure

cell signaling-digestion

Trypsin: breaks down proteins during digestion. called PROTEASES
Amylase: starch breaker and secreted by pancreas

cell signaling-metabolism

Alcohol dehydrogenase: helps metabolise ethanol.
1cza Hexokinase: adds phosphate to glucose, after glucose is taken up by cell, they usually add phosphates to molecules.

Oxygren transport: Metabolism

Hemoglobin, binds to oxygen in lungs and carries it in blood to tissues for use in metabolism.

Metabolism and energetics-Membrane proteins

Atp synthase: generates ATP for use in cellular function

Immune protection

Antibodies bind to cellular invaders like bacteria and viruses to help protect the body from infection.

Replication and maintenance signalling

DNA polymerase: Binds to one strand of DNA and adds the complimentary strand to it
RNA polymerase: creates a single strand of RNA that is complementary to one of the strands of duplex DNA.

Amino acids are

Chiral

L-form- Co group on left

D-form- co group on right

amino acid zwitterion

Amino acid facts

20 amino acids have common backbone but different side chains.

Non polar side chains amino acids

These side chains on amino acids will be mostly hydrophobic (insoluble in water). No charged or electronegative atoms are present to form H bonds

Top row: Help drive protein folding, found inside protein.

Bottom row: Larger hydrophobic, have larger sidechains. They absorb ultraviolet light, used in protein spectroscopy.

Glycine, Gly, G

Not chiral, flexible, almost in a group by itself

Proline, Pro, P

Sidechain binds to amino groups, IMINO acid, rigid, almost in a group by itself

polar side chains Charged

Divided into acidic and basic.
Acidic: aspartate(D) Asp
Glutamate (e) Glu

Basic: Lysine (K)
arginine (R)
Histidine(h)

Difference between aspartate and glutamate

Aspartate has 4 C
Glutamate has 5 C

Where is histidine normally found

active sites

Polar sidechain uncharged

serine, threonine, cysteine, tyrosine, asparagine, glutamate.
All of them can participate in hydrogen bonds

Serine

found in active sites, involved in catalyzing protease reaction

Threonine and asparagine

known to be glycosylated in mammalians/to improve solubility of protein, to lead them in correct sites

Cysteine

good Nucleophile, can be involved in disulphide bonds in an oxidising environment. in reducing environment they can split up

Tyrosine

susceptible to phosphorilation, turn enzyme on/off.
It has an amromatic sidechain it can absorb ultraviolet reactions.

One letter abbreviation

E6V
-1st: original or native amino acid
-Number= location of mutation of protein
-Second letter: new or mutated residue.

How can ionisable sidechains be classified

by their pka value

pKa

value for an amino acid or protein is the pH at which the group is 50% ionised

pI, isoelectric point

ph at which the net charge on an amino acid/protein is ZERO

Amino acids that have ionisable side chains

Amino acids that have ionisable side chains 2

cysteine is a good nucleophile. pka 8
'Tyrosine has hydroxide group pka 10 (basic)

Pka values for amino acids

Modified amino acids are

translated from RNA proteins at ribosome, some are modified after they are added to proteins, post translational modification.

what type of bond can a cysteine form with another nearby cysteine

Covalent bond which is called the disulphide bond

Examples of amino acid modification

Phosphorylation
Hydroxylation
Carboxylation
Metal binding
iodination
Glycosylation

Phosphorylation

Used to control enzyme activity

Hydroxylation

needed to prevent connective tissues diseases and scurvy

Carboxylation

needed for blood clotting, often glutamate involved

Phosphoserine

Glycosylation of threonine

Asparagine can often be glycosylated as well. Haemoglobin also which is used to diagnose and follow patients with diabetes

Peptide bond

covalent bond formed between amino acids

Dehydration reaction

A chemical reaction in which molecules combine by removing water

Peptide bond characteristics

-40% double bond character leads to planarity. planar conformation maximizes pie bond overlap.
-Rotational barrier of ~80kJ/mol
-Peptide bond is predominantly trans.

PLANAR, TRANS, DIPOLE

Methaionine 184, met 184, M184

Methionine residue at position 184 in a protein or peptide

Description of protein shape

Mostly globular so amin chain is to form a more compact shape and be able to double back.
Globular proteins will turn out to be comprised of primarily alpha helix, beta structure and turns.

Levels of protein structure

1. Primary- amino acid sequence of protein
2. Secondary-local 3d arrangement of protein chain over short stretch of adjacent amino acid residues
3. Tertiary-3D structure of a complete protein chain
4. Quaternary-interchain packing and structure for a protein that contains multiple protein chains

primary protein structure

sequence of amino acids

secondary protein structure

coiling or folding of a polypeptide due to H-bonding between amino acids

Tertiary protein structure

3D folding pattern of a protein due to side chain interactions
MYOGLOBIN

quarternary protein structure

A number of polypeptide chains linked together, and sometimes associated with non-protein groups to form a protein.

What are the main chain atoms of a protein

N, C alpha, C'

Name of bond angle between N and C'

omega (W) very clost to 180 0r 0

Name of bond angle between N and C alpha

phi(ϕ) 0 to +/- 180

Name of bond angle between C alpha and C'

psi(Ψ) 0 to +/- 180

Protein structure contain many _________ bonds

flexible

phi and psi bond angles

N and C alpha phi
C alpha and C' PSI

Phi-Psi Restrictions

-Phi rotation can lead to O-O collisions
-Psi rotation can lead to NH-NH collisions
due to STERIC HINDRANCE

Peptide bonds can be trans or cis

Most are trans.

trans peptide bond

W is about 180 and C alpha atoms are found on opposite sides of the peptide bond

Cis peptide bond

atoms found on same side of the peptide bond, w is about 0 degrees.
Steric hindranc eis increased for cis peptide bonds

protein secondary structure

Dominated by two canonical structures.
B strand
B sheet

In the alpha-helix, the main chain spirals around the central axis like a spiral staircase. RIGHT HANDED spiral.

What are non covalent interactions in helices called

Hydrogen bond, between the carbonyl residue "n" and n-h residue of "n+4"; o-n distance ~2.9A

What do hydrogen bonds help do

stabilise the alpha helix structure

Amide hydrogens

Have a partial positive charge

Carbonyl oxygens

have partial negative charge

Key properties of alpha helix

-Spiral is right handed
-Sidechains point out from the helix axis; help stabilise helix.
-Stabilising hydrogen bonds, 3-7kcal/mol or 12-28
phi=~-57 deg
psi=~-47 deg
-Some residues are helix breakers like glycine and proline
-Helix dipole exist,~positive at N terminus

Helix dipole depiction

Helix wheels

Help illustrate helix side chain arrangements .

3.6 residues/turn
One full tuen is 360 deg.
IN an alpha helix, each amino acid side chain is seperated by 100 deg

Beta structure

-Comprises of stretches of residues called B-strands
-Hydrogen bonding occurs between adjacent chains (strands)
-Adjacent chains can often form a B-sheet> 2 B-strands
-2 to 10 strands per sheet
-Average strand length contains ~6 amino acid residues and each strand contain up to 15 residues.
-2 types of hydrogen bonding interaction in B-sheet

What are the 2 types of hydrogen bonding interaction in a B-sheet

Parrallel and antiparallel

Finding the direction of the strand

Find amide nitrogen and look for alpha carbon, direction is from N to C alpha

B-structure properties

-Extended, pleated, 'B-pleated sheet"
-"Sheet" versus " strand"
-Sheets not planar, r-handed twost
-Side chains point above and below sidechains.
-Any NP-P-NP-P stretch of residues commonly form a B-strand

beta structure in silk

Different physical properties, it is antiparrallel strand. very flexible

Turns: key properties

Needed to form globular proteins
-often short, involve 3-4 residues
-30% of residues involved in turns.
-High gly, pro content
-h-bond across turn is common
-type 1 and 2 are common.

Consensus protein structural display

-helices shown as spiral/cylinders
-Strands shown as arrows, pointing from N to C.
-Turns and random coil, shown as loops or rope like stretches

Protein structure shorthand

Ribonuclease A structure shorthand

Protein structure shorthand advantages

Easily visualioze main path of protein
-identify elements of secondary structure
-comparison

Lecture6

Common motifs of supersecondary structure

helix-turn-helix,
beta hairpin,
greek key,
strand-helix-strand

helix-turn-helix

most common DNA binding motif (dimmers). Also present with loop in calcium binding proteins

b-hairpin

-common
-antiparallel
-length vaires

Greek key

4 antiparallel strands (strands 1,2 in the middle, 3 and 4 on the outer edges)

Strand, helix, strand

arrow, strand, arrow

what combines to form domains

supersecondary structure elements

Domain

independently folded regions that often possess a specific function within the protein

Protein domain

has a hydrophobic core and the hydrophilic parts of the protein are arranged on the surface in contact or near solvent.

Hydrophobic cores

Important for protein stability

Name the 3 domain families based on tertiary structure

-alpha domain family
-alpha/beta family
-antiparrallel b family

Alpha domain family

mostly helical

alpha domain family globin fold

Amphipathic helices with side chains closely packed together within hydrophobic core.
Packing can occur between nonadajacent helices

alpha/beta family

mix of alpha and beta structure

Alpha/Beta Family

The ones sticking out of the barrel are all polar or charged

anti-parallel beta family

Mostly antiparallel structure

Protein folding

The physical process by which a polypeptide folds into its characteristic three-dimensional structure, which is essential to the protein's function.

The Anfinsen experiment

The hypothesis that "protein amino acid sequence determines the final shape a protein assumes in a water solution" was proven to be correct when Christian B. Anfinsen showed that if the enzyme ribonuclease was opened out into a linear chain and then allowed to reform, it reassumed the correct catalytic shape.

Folding pathways

1. formation of short sec structure segments.
2. subdomains forms
3. Subdomains come together to form a partly folded domain; a molten globule that can rearrange.
5. final domain structure emerges, small conformational adjustments to give final compact native structure

Stabilisation of protein folding

• Non-covalent interactions, while individually weak in proteins, collectively make a significant contribution to protein conformational stability
• In some proteins additional covalent bonds (eg. disulfide bonds) may be present that contribute to conformation stability
• The hydrophobic core is likely the most important noncovalent contributor to protein stability in aqueous solution.

What is the most important noncovalent contributor to protein stability in aqueous solution

Hydrophobic core

some protein folding is assisted by

chaperones

A. Chaperone-independent
b. Chaperone-dependent
c. Chaperone dependent

What do chaperones do

The are responsible for the correct folding of proteins and help proteins reach their correct tertiary structure.