BIO 2B03 TEST 1 MODULES 1-4 + Applied Lectures (INCOMPLETE - NO TIME TO FINISH THIS QUIZLET LOL)

What is a protein?

A string of amino acids (a polypeptide chain or polymer of other small molecules)

What are some of the functions of proteins?

- Structural components of the cell

- They are the signals produced by cells

- They are the receptors that receive signals

- Sensors for environmental change

- Enzymes, catalysts for chemical reactions

- gene regulation

True or False: There is a virtually unlimited amount of protein structures that proteins can form, unlike RNA and DNA

TRUE

Each protein has its own unique _______________ that enables it to carry out a specific_________________?

Each protein has its own unique STRUCTURE/FORM that enables it to carry out a specific FUNCTION

When amino acids are attached in a linear array, what protein structure does this create?

This creates the primary structure of the protein which can also be called a polypeptide!

Peptide Bond

formed by a condensation reaction (water released) between the amino group of one amino acid and the carboxyl group of another

N-terminus

The end of a polypeptide chain or protein that has an amino group

C-terminus

The end of a polypeptide chain or protein that has a carboxyl group

How many amino acids are there?

20 different amino acids

What is the basic structure of all 20 amino acids?

A central alpha carbon, with 4 side chains

1. Amino group

2. Carboxyl group

3. Hydrogen

4. R-Group

True or False: It is the variable R group that will define the property of each amino acid residue, and it is the ACCUMULATED R groups that will define the properties of a polypeptide or protein

TRUE

How can R-groups differ?

- Size

- Shape

- Charge

- hydrophobicity

- reactivity

Amino acids are classified into groups based upon?

Based upon either their solubility in water or the polarity of the side chain

Hydrophilic Molecule

One that is typically charge polarized and capable of hydrogen bonding with water

Hydrophobic molecule

One that is not electrically polarized and unable to form hydrogen bonds, thus water repels them in favour of bonding with itself

What kinds of molecules are typically hydrophobic?

Oils, fats - saturated hydrocarbons (long carbon chains linked together by single bonds)

- The general formula is CnH2n + 2 == alkane

True or False: Hydrophobic amino acids tend to be found in the interior of soluble proteins, and they form a hydrophobic core

TRUE - a protein that is found in a hydrophobic environment such as the membrane, will have the opposite structure, with the hydrophobic amino acids accumulating on the exterior of the protein

How are the hydrophobic amino acids divided up into 2 categories?

1. Aromatic amino acids (those with aromatic rings)

- Phenylalanine, Tryptophan, Tyrosine (does have OH group rendering it also slightly soluble)

2. Aliphatic amino acids (those with long carbon chains)

- Alanine, valine, isoleucine, Leucine, Methionine

Where would you normally tend to find hydrophillic amino acids of SOLUBLE cytosolic proteins?

On the exterior of soluble cytosolic proteins

Hydrophillic amino acids are also grouped into 2 categories, either being charged, or polar and uncharged. What are the charged amino acids?

Basic Amino acids: Positively Charged

- Lysine, Arginine

Acidic Amino Acids: Negatively Charged

- Aspartic acid, glutamic acid

What are the Polar, and uncharged amino acids?

Serine, Threonine

- uncharged at neutral pH but have polar -OH groups that participate in H-bonding

Asparagine and Glutamine

- Uncharged, but have polar amide groups

Other amino acids do not fit into being only hydrophillic or hydrophobic, they are special, what are these amino acids?

Cysteine: able to form covalent bonds with other cysteine residues which are called disulphide bridges or cysteine bridges

Glycine: small, allows bends in the polypeptide chain

Proline: Side chain forms a bond with the amide side chain of the amino acid, producing a kink in the peptide chain, essential for structure of many proteins

Histidine: has amino diethyl side chain that shifts between positive charge and a neutral charge depending on pH of environment

Which terminus are amino acids added to?

Amino acids are added to the C-terminus (carboxyl end)

What does the small subunit of the ribosome do in translation?

The small subunit of the ribosome positions mRNA so it can be read in groups of 3 letters known as codons

What does the large subunit of the ribosome do in translation?

The large subunit removes each amino acid and joins it on the growing protein chain

What are the 3 sites on the ribosome, and what occurs at each site?

A site: tRNA enters the ribosome and is tested for a codon - the anticodon (on tRNA) matches with the mRNA

P site: provided there is a match, tRNA is shifted to P-site and the amino acid it carries is added to the end of the amino acid chain

E site: Spent tRNA is moved to E site and is ejected from the ribosome to be recycled

What are the 4 levels of protein organization?

primary structure (linear arrangement of amino acids), secondary structure, tertiary structure, quaternary structure

The number of different polypeptide sequences (primary sequences) is limited by what factors?

1. 20 distinct amino acids that can become incorporated into a polypeptide

2. The number of amino acids found in the polypeptide

For any polypeptide chain that is n amino acids long, the number of different arrangements of amino acids in the chain would be defined by 20^n

How many different polypeptide chain can be produced from 4 amino acids

20x20x20x20 amino acid sequences can be produced

160 000 total sequences for 4 amino acids

True or False: Polypeptides fold spontaneously and assume a random-coil structure

TRUE

What does the term statistical coil mean?

Proteins might not have a single stable structure, but a collection of related structures they switch between

- the term is a representation of this idea and suggests that the protein spends most time in a particular structure, but not 100% of the time

Native Structure of a protein

The structure that the polypeptide assumes most of the time

What types of chemical interactions stabilize interactions of polypeptides?

Ionic bonding

Hydrogen bonding

Van der Waal forces

Hydrophobic effects

True or False: Specific amino acid identity does not affect folding

TRUE - it is more to do with the interactions between amino acids and their characteristics that affect folding

Secondary Structures (proteins)

Periodic folding of the polypeptide chain into distinct, conserved, geometric arrangements (alpha-helix, Beta sheets, turns, loops)

What are secondary structures of proteins defined by?

The patterns of hydrogen bond formation that occur between the non-variable amino acid side chains and the amino and carboxyl groups

Peptide bonds connect neighbouring amino acid residues, however in the alpha helix, what forms the shape?

In the alpha helix, every amino acid residue forms a hydrogen bond with an amino acid residue 4 positions away

It is the periodicity of hydrogen bonds that define the alpha helix structure

Are alpha helices found in all proteins, in the same pattern and shape and structure? If so, how is this possible?

Yes these alpha helices are found in the exact same pattern and same shape and structure in all proteins

This is due to the fact that the VARIABLE R GROUPS ARE NOT INVOLVED IN THE FORMATION OF THESE STRUCTURES

What do the R-groups in the Alpha helix determine?

The R-groups in the Alpha helix determine the hydrophobic/hydrophillic quality of the outer surface of the helix

beta-pleated sheet (β-pleated)

secondary structure found in proteins in which "pleats" are formed by hydrogen bonding between atoms on the backbone of the polypeptide chain

Are the positions of the hydrogen bonds in beta pleated sheets regular like the hydrogen bonds in alpha helices?

No, the length of the beta strands vary

- Beta strands may be aligned anti-parallel, or in a parallel arrangement in which neighbouring strands are both aligned in the same orientation from N-terminus to C-terminus

Do R groups contribute to the formation of the structure of beta-pleated sheets?

No, the R-groups don't contribute to the formation of the structure, they are only important for their properties - side chains point top and bottom

Hinges, turns, loops

Ways of describing the bends in the polypeptide backbone

An example of a conserved connector is the beta turn, what is the beta turn?

The Beta turn involves 3 or 4 amino acid residues and is often found connecting the strands of a beta-sheet

What are Motifs?

unique collections of secondary structures where specific amino acid residues are required to maintain the conserved structure of the motif

Motifs are often associated with a particular function

Coiled-coil motif

Two alpha helices wrapped around one another

How is a coiled-coil able to be formed?

Due to the fact that the alpha helices are amphipathic, with both hydrophillic and hydrophobic suefaces

In an aqueous environment like the cytosol, hydrophobic effects favour the association of the hydrophobic surfaces, which holds the helices together

How are the continuous hydrophobic surfaces along the alpha helices formed?

In order to form these hydrophobic surfaces, the hydrophobic amino acid residues must be at positions 1 and 4 within a heptad repeat (a repeat of 7 amino acid residues)

leucine zipper

A simple version of a heptad repeat which shows the pattern LEU-X6-LEU-X6-LEUX6 where X6 is any 6 amino acid residues

Where is the coiled-coil motif most commonly found?

It is most commonly found in the DNA binding proteins because the dimensions and properties of this structure allow it to fit within the grooves of the double stranded DNA helix

What is the Zinc-finger motif?

Consists of on alpha helix and 2 bets strands (small beta sheet), held in position by the interaction of precisely positioned Cyc(c) or His(H) residues with a zinc atom

where are zinc fingers found

These motifs are most often found within DNA binding proteins, but are also able to bind to RNA molecules as well

B-Barrel Motif

A beta sheet forms a barrel when the last beta strand forms hydrogen bonds with the first strand

- Essentially a large beta sheet that loops back upon itself

- Collection of 4-10 anti parallel beta strands form a sheet

What is the beta barrel motif useful for?

Useful for forming a channel or a pore across a hydrophobic membrane

Structure would be amphipathic - exterior of barrel is hydrophobic whereas interior is hydrophillic

Helix-loop-helix motif

Two small alpha helices joined together by a loop region

- Loop region can bind to calcium via the carboxyl side chains from Asp or Glu in the loop

True or False: the shape of the helix-loop-helix motif can only be established once the polypeptide is interacting with the calcium

TRUE - protein structure and function are therefore dependent upon the cofactor

Tertiary structure (proteins)

Three dimensional arrangement of all amino acid residues of a single polypeptide

The fundamental unit of the tertiary structure of a protein is the domain, what is a domain?

A domain is a substructure produced by any part of a polypeptide chain that can fold independently into a compact, stable structure

- Often a functional unit

This allows researchers to study domain function independently of the whole protein, but then consider how the protein acts as a whole

Functional Domains (proteins)

Regions of a protein that perform a certain activity

e.g. DNA binding, enzymatic, protein-protein interaction

Structural domains (proteins)

regions of protein that form compact, largely independent globular domains

e.g. proline-rich, acidic domain

Src protein

an important protein in the regulation of the cell cycle

How many domains are present on the Src protein?

2 functional domains: Small kinase and large kinase domain

2 structural domains

1. SH2 domain (Src homology 2 domain) - this domain does have an identified function, recognizing phosphotyrosine residues in other proteins

2. SH3 domain (Src homology 3 domain)

Quarternary Structure (protein)

Assembly of a multimeric protein

- multimeric protein == a functional protein composed of multiple folded polypeptides

Dimer

2 polypeptides or two subunits

Trimer

3 polypeptides

Homodimer

Two identical polypeptides

Heterodimer

Two different polypeptides

Intrinsically unstructured proteins

Proteins that lack a tertiary structure as isolated subunits

Bioinformatics studies predict that a significant fraction of the genome codes for unstructured proteins, and that the fraction increases with the complexity of the genome

True or False: We might also find that unstructured proteins will assume specific folding patterns when they are interacting with their substrate

TRUE

post-translational modification

Alterations of the chemical properties of the variable R-groups which change protein structure and function

Acetylation

The addition of an acetyl group

- this modification has been found to occur on many proteins where it acts to protect proteins from proteases

Methylation

a biochemical process that influences behavior by suppressing gene activity and expression through the addition of methyl groups

True or False: Both Acetylation and Methylation are reversible

TRUE

Phosphorylation

Transfer of a phosphate group from ATP to the -OH group of SERINE, TYROSINE, or THREONINE by kinases

Dephosphorylation to remove phosphate groups is catalyzed by enzymes called?

Phosphatases

What is the function of phosphorylation?

Both Phosphorylation and dephosphorylation can activate and deactivate proteins by changing the shape of the protein or by changing the ability of the protein to interact with a substrate

Hydroxylation

Addition of a hydroxyl group

- important for changing the structure of proteins

The enzyme that catalyzes hydroxylation in the cell requires what cofactor?

Vitamin C

Carboxylation

Addition of carboxyl group (COO_)

- changes the properties of the amino acid residues by adding a negative charge

Can facilitate ion bond formation or allow a positively charged cofactor to bind

Glycosylation

- Addition of carbohydrates

- Sugars are added to -OH groups of SERINE AND THREONINE

Glycosylation is important for protecting proteins from proteolysis and for proper protein folding

Lipidation

Addition of lipid molecules

- important for anchoring proteins to hydrophobic biomembranes

What are the three foundational pillars of protein folding

- It is spontaneous

- It is reversible

- It is unique

True or False: the process of protein folding is not targeted, it is simply that stable states are maintained for longer periods of time once they occur

TRUE

Hemoglobin

Tetramer containing 4 subunits (2 alpha subunits and 2 beta subunits) that transports oxygen in the blood

In people with SCA, what occurs?

Single amino acid change in beta globin gene sequence which substitutes glutamate to valine at position 6 - this makes the protein more hydrophobic, as valine has less charge than glutamate, and causes aggregates

Alpha-1-antitrypsin

Protein found in the cells of the lung important for maintaining elasticity

What amino acid change causes emphysema?

Aspartic acid at position 256 is replaced with valine, there is a change from a charged amino acid reside to a hydrophobic amino acid residue

- this changes the primary sequence, which affects the tertiary sequence of the protein

What are some of the ways in which normal proteins can break down?

1. Genetic amyloid related diseases: single changes in nucleotides change A.A sequences and lead to unstable intermediates that can form fibrils (Aggregates)

2. Peptide Fragments of a protein that are not folded into the stable globular whole - fibrils form

3. Breakdown of chaperone system to prevent intermediates that tend to result in fibril formation

4. Infection by misfolded fibril structures

Prion Diseases

Fatal transmissible neurodegenerative diseases

- The results of the disease are neuronal cell death in the brain

Why are Prion diseases so dangerous?

Because of the length of the incubation period where there is no conventional immune response that can alarm that anything wrong is taking place

How do Prion Diseases arise?

Modes of acquisition

- Spontaneous

- Heritable

- Infectious

What are Prions?

Prions are an abnormal form of a normal protein

The body contains normal cellular versions of the Prion protein

What are the differences between the normal versions of the prion protein (PrPc) and misfolded versions of the protein (PrPSc)?

PrPc - normally soluble, sensitive to protenases, do not aggregate, rich in alpha helices

PrPSc - Rich in beta-pleated sheets, insoluble, protenase resistant, tend to aggregate

What gene codes for prion proteins in Humans?

a gene called PRNP

What are the major clinical features of prion diseases

Progressive neurological decline

- Rapidly progressive dementia

- Ataxia/uncoordinated movements

- Visual problems

- Akinetic mutism - inability to move or speak

Cellular Prion Protien PrPc

A glycosylphosphatidlyinositol (GPI) anchored membrane glycoprotein

- Normally expressed in many different cell types, with elevated levels in neuronal synaptic membranes

Can sporatic misfolding of normal cellular prion proteins result in Prion disease? What happens when this occurs ?

Yes it can result in prion disease and when this occurs, the prion proteins transition from alpha helix rich proteins to proteins rich in Beta sheet secondary structures

- Beta sheet rich proteins have higher tendency to aggregate

The misfolding of these prion protiens can also induce other existing PrPc proteins to convert into the associated misfolded form of the protein - INFECTIOUS

This leads to eventual formation of insoluble, fibrous protein aggregates

What is the suspected mode of infection of Prion Proteins?

The suspected mode of infection involves the instability of transient protein states

While PrPc proteins are in transient folding states, Beta sheet rich versions of the prion proteins are better able to influence the misfolding of the normal proteins into the beta sheet rich and misfolded prion forms

THE END RESULT: PrPSc proteins are now able to nucleate or build the formation of amyloid fibrils

These fibrils break into multiple new "seeds" where these misfolded proteins can go on to continue the conversion to abnormal PrPSc proteins

Where are PrPC proteins synthesized?

They are synthesized, folded and glycosylated in the ER

- Two fatty acids in the GPI anchor the protein to the cell membrane

PrPSc accumulates near PrPC leading to potential for conversion

Conversion can also occur in endosomes or lysosomes

What would an amino fluorescent staining of both infected and uninfected cells look like (PrPc)?

UNINFECTED CELLS: normal PrPc proteins are localized mostly to cell membrane

INFECTED CELLS: cell surface signals of normal PrPc proteins is decreased in cells that have been infected with prions - PrPc proteins seem trapped in intracellular compartments