PHR911: Block 2 Lecture 2 Amide Bonds and Protein Structure

Primary Protein Structure:

Order of amino acids

What kind of bonds do primary protein structure have?

Covalent (amide) bonds

Hydrolysis:

Addition of water

Condensation: 

Loss of water

Conservative proteins:

If an amino acid is substituted by another amino acid of the same classification

Non-Conservative Protein:

A missense mutation where one amino acid is replaced by an amino acid not of the same classification

Variable vs Invariable:

If amino acid changes are tolerated or alter the function of the protein

Humalog:

Engineered insulin with B28 and B29 swapped

How was Lantus synthesized?

Recombinant DNA technology using E.coli

Lantus:

GlyA21

ArgB21

ArgB32

Human insulin

Gene:

Piece of DNA that encodes for a functrional protein

Allele:

One of a number of alternative forms of the same gene

No consequence variation of primary structure:

Polymorphism: Varients of an allele that occur with significant frequency

Detrimental variations of primary structure:

• Lead to a defective protein

An example of detrimental variations of primary structure:

Lactose intolerance

Beneficial variations of primary structure:

Leading to superior function/activity

Isoforms (isozymes):

Protein with variations in primary structure but have essentially the same function

An example of isoforms in humans?

Adenylate cyclase- at least 9 different isoforms.

Developmental variation in primary structure example:

Fetal and embryonic hemoglobin has higher affinity for O2 than in adult hemoglobin.

Protein paralogs:

Groups of proteins with similar (but not identical) structure and function that have evolved from the same gene following duplication

Example of a paralog:

Myoglobin and hemoglobin

Posttranslational modification:

Occurs after the primary structure of a protein is made (translation)

Why does post-translational modification occur?

• Regulation

• Anchor protein in membrane

• Enchance protein-protein interactions

• Target protein for degradation

Glycosylation: Modified AA

Process of attaching sugar molecules to proteins.

N-linked or O-linked sugars

Often found on extracellular side of plasma membrane.

Fatty acid acylation or prenylations: Modified AA

Covalent attachment of fatty acids to a protein

Increases hydrophobicity

Modified AA: Kinase/Phosphatases:

Ater protein activity and regulatory function

Oxidation: Modified AA

Loosing an electron

Carboxylation:

Addition of a COO-

Simple proteins:

Only contain AA

Conjugated proteins:

Contain components other than amino acids

Glycoproteins:

Contain sugar

Lipoproteins:

Contain lipids

Metalloproteins:

Contain metal ions

Nucleoproteins:

Bind nucleic acids

Hemoproteins:

Contain a heme group

What isomer is thermodynamically favored?

Trans

Residue:

Amino acid in a protein

Peptide sequences always begin at the what?

N-Terminus

Forces stabilizing protein structure:

Hydrogen bonding

Hydrophobic effects

Electrostatic interactions

Disulfide bonds

Van der waals

Secondary Structure:

Recurring, localized structure as a result of order of amino acids

Most common secondary structures:

Alpha-Helix

Beta-helix

Alpha-Helix:

A carbonyl oxygen makes a hydrogen bond to every n+4 amide hydrogen

A rigid, stable conformation maximizing hydrogen bonds while maintaing allowed torsion angles

What isomer are alpha helix side chains

Trans

What AA is not found in alpha-helix?

Proline

Beta Sheet:

Maximizes hydrogen bonds while maintaining allowed torsion angles

Carbonyl oxygen is hydrogen bonded to amide nitrogen of a different strand

Beta-Sheet Parallel:

2:1 amino acid hydrogen bonding

Tend to have hydrophobic R side chains on both sides of the sheet

Beta Sheet: Antiparallel

1:1 AA hydrogen bonding

Tend to have hydrophilic side chains on one side, hydrophobic on the other

What do beta-turns often contain:

Asn-Gly

Pro-Gly

Why does beta turns have high amounts of Gly residues?

Their simple side chain of H allows for conformational flexibility and limited steric hinderance.

Beta sheets tend to twist towards the right or left?

Right

What is the overall shape of protein?

Tertiary Structure

What determines the function of a protein?

Position of the amino acid side chains in a 3D space

The 3D structure of a protein must have a what for a specific molecule?

Binding site

The external surface of a 3D protein must have what conformation?

Stable Conformation

A 3D protein structure must be able to be what when it is damaged/no longer needed?

Degraged

Globular Proteins:

Usually water soluble, compact, roughly sperical

Hydrophobic interior, hydophilic surface

What is the core of a globular protein?

Nonpolar side chains (hydrophobic effect) and polar, uncharged amino acids (h-bonds)

What is the peripheral outside of a globular protein?

Charged and polar amino acids

Fibrous Proteins:

Assembled into long cables or threads

What do fibrous proteins do?

Offer mechanical support

Examples of fibrous proteins?

alpha-keratins: Major components of hair and nails

Collagen: major component of tendons, skin, bones, and teeth

What are membrane proteins?

Consist of hydrophobic regions and hydrophilic regions.

What do membrane proteins do?

Signal transduction

ion channels

porins

receptors

Beta₂ - adrenergic receptor:

Adrenaline

What are transmembrane helices predicted by?

Hydrophobic stretches of 20-25 aa residues

GPCRs:

G-protein coupled receptors

Real important class of proteins mediating communication between cells, tissues, and organs and operating signal transduction pathways

Characteristics of tertiary structure in membrane proteins:

Posttranslational modifications

Conformational changes → signal transmission

Motifs:

Common elements of secondary structure seen in many polypeptides

• Typically contains alpha-helix and/or beta-pleated sheet

Domains:

Functional, independent regions of a poypeptide defined by a tertiary structure

• Transmembrane domains

Quaternary Structure:

Individual proteins (subunits) that fulfill their function together

Dimers:

2 quaternary proteins

Trimers:

3 quaternary structures

Tetramers:

4 quaternary structutres

What are some advantages of having quaternary structure?

Increase protein stability

Alter binding affinity for ligands (cooperativity, increased affinity)

Different subunits can have different activities that cooperate in a different function

Homo:

2 or more of the SAME protein

Hetero:

2 or more of DIFFERENT proteins

Structure of proteins determines what?

Function

What are common targets for drugs?

Protein receptors

Enzymes

Small molecules that bind to protein receptors:

Ligands

Binding affinity:

One ligand (L) binding to one protein (P) to form a complex (LP)

The greater the K_a the greater the affinity of the what?

Protein for a ligand

When you have a higher affinity ligand, what is the K_d  

Smaller K_d  value

Myoglobin and Hemoglobin tertiary structures are entirely what?

Entirely an alpha-helix

What doe myoglobin and hemoglobin do?

Oxygen binding proteins with similar primary structure

How many oxygen binding sits does hemoglobin / myoblobin have?

4

Does myoglobin or hemoglobin have a higher affinity to oxygen?

Myoglobin

Prosthetic group of myoglobin/hemoglobin:

Small organic molecule that binds tightly to protein and its integral for the function of a protein.

Structure of prosthetic group in myoglobin/hemoglobin:

Ferrous iron binding 4 pyrrole groups

HIS side chain

Ozygen

What happens with oxygen binds to myoglobin/hemoglobin?

Oxygen binding changes the conformation of the protein, affecting quaternary structure of hemoglobin

Hemoglobin and Oxygen binding is what kind of cooperativity?

Positive

The binding of 1 oxygen increases the rate of what?

Binding of the next oxygen

Immunoglobins: Binding protein:

Antibodies bind foreign ligands called antigens or epitopes to initiate inactivation or destruction of a foreign object

What is the key ingredient in ELISAs testing kits?

Antibodies

Native protein folding confirmation:

Every molecule of the same protein folds into the same tertiary structure

Protein Denaturation:

Change in the shape of protein, usually causing loss of function.

What can cause protein denaturation?

pH changes

Temperature

Salt concentration

Misfolding:

Either loss of function or often aggregate

Protein aggregates are often insoluble and what kind of illness?

Prion-related

Alzheimers:

Amyloid protein misfolding

Mad cow disease and Creutzfeldt-Jakob disease:

Prion protein misfolding