Chapter 5

Question 1

What is a ligand?

Answer 1

Any molecule that interacts with the binding site of of a protein. Does not covalently bind, so it is reversible! (ligand can come back out)

Question 2

What is a substrate?

Answer 2

Something that an enzyme acts on

Question 3

Where do disulfide bonds form and why? How do they affect protein structure?

Answer 3

Disulfide (covalent) bonds are formed between the sulfur atoms of two cysteine residues. These bonds play a crucial role in stabilizing the tertiary and quaternary structures of proteins.

Question 4

What is meant by “reversible binding” of a ligand to a protein?

Answer 4

Ligands can be reused after detaching from the protein once the desired reaction occurs.

Question 5

What aspect of thermodynamics drives protein folding?

Answer 5

ENTROPY: Because the water is more ordered when the protein is unfolded, it would be favourable to fold the protein because after folding then the water molecules can move more freely –> higher entropy –>negative ΔG –> spontaneous folding.

Question 6

What is a prion and what does it do?

Answer 6

An unusual and infectious agent composed primarily of protein material. Unlike viruses, bacteria, or fungi, prions do not contain genetic material like DNA or RNA. PRIONS CAUSE PROTEINS TO MISFOLD into an abnormal three-dimensional structure, causing major problems in a short amount of time.

Question 7

Compare and contrast chaperone proteins and chaperonins. What do they do?

Answer 7

Chaperones: proteins that assist the conformational folding or unfolding of large proteins or macromolecular protein complexes.

Chaperonins: a type of chaperone protein

Question 8

What is a the difference between Nonspecific Interaction and Specific Protein Binding?

Answer 8

Nonspecific Interactions: Unspecific, weaker forces. Electrostatic interactions (between charges), hydrogen bonds,
and hydrophobic interactions play a role
(higher Kd, lower affinity)
* EX: Water with anything

Specific Protein Binding: between protein and specific target molecule or ligand. Tight Hydrogen bonds result in DNA bending. Many DNA binding proteins interact in the major groove! (lower Kd, higher affinity)
EX: antibodies with antigens

Question 9

What is an operon?

Answer 9

A group of genes that are arranged together in a specific order on a DNA strand. These genes work together to carry out a particular function in a cell. What makes an operon special is that these genes are all controlled as a single unit and are turned on or off together.

Question 10

What are promoter and operator sequences?

Answer 10

Promoter Sequence:
Starting point for reading the genes. It’s a signal that tells the cell’s machinery to begin making a copy of the genes.

Operator Sequence:
A region near the genes that can be turned on or off. Regulatory proteins can attach to the operator and control whether the genes are actively being read or not.

Question 11

What is a repressor?

Answer 11

A regulatory protein that binds to an operator sequence and prevents the transcription of specific genes near that sequence.

Question 12

What is the charge of DNA?

Answer 12

Negative (phosphate backbone is negative in solution)

Question 13

What is the Dissociation Constant (Kd)? What does it tell us?

Answer 13

Where half the available ligand binding sites are occupied (think of a graph)

Kd tells us the strength of the binding that will occur

Question 14

What does a high or low Kd value tell us?

Answer 14

• Higher Kd means lower affinity
• Smaller (higher negative exponent) number for Kd
  means tighter binding.
• SUPER HIGH AFFINITY INTERACTIONS (really tiny Kds) MAY BE CONSIDERED IRREVERSIBLE

Question 15

What are two factors that play a role in binding affinity?

Answer 15

1. Chemical Complementarity
2. Specificity

Question 16

Why do some proteins have high affinities for their ligands while others do not?

Answer 16

Chemical complementarity (compatibility) with different shapes, sizes and chemical properties plays a huge role in binding affinities, as well as the rate at which proteins are able to associate with specific ligands.

Question 17

Compare and contrast the major and minor groove in DNA. How, where, and why do proteins usually bind DNA?

Answer 17

The major groove occurs where the backbones are far apart, the minor groove occurs where they are close together.

How: certain proteins bind to specific sequences

Where: in either of the grooves

Why: to regulate, replicate and repair genes

Question 18

How does the Lac repressor regulate the lac operon?

Answer 18

The Lac repressor binds to two operators to shut down transcription via a loop by preventing RNA Polymerase to bind to the promoter.

Question 19

Why does DNA polymerase need magnesium ions?

Answer 19

Magnesium acts as a “cofactor” for DNA polymerase

Question 20

What are cofactors?
Provide examples.

Answer 20

INORGANIC compounds (such as non-protein chemical compounds or metallic ions) that are required for an enzyme’s role as a catalyst (3RD PARTY)

ex: magnesium is needed at the active site to neutralize the negative charges in DNA. (DNA is negative, cofactors bear a positive charge)

Question 21

What are coenzymes. Provide examples.

Answer 21

SUBSET OF COFACTORS

ORGANIC compounds required by many enzymes for catalytic activity (3RD PARTY)

ex: Acetyl-Coa and vitamins

Question 22

How does the enzyme structure catalyze reactions?

Answer 22

Enzymes have specific complementary structures to their substrate. This allows them to “strain” the substrate moving them into the transition state, which provides the catalytic properties as they lower the activation energy.

Question 23

What is a retrovirus?

Answer 23

A type of virus that inserts a DNA copy of its RNA genome into the DNA of a host cell that it invades, thus changing the genome of that cell

• USING REVERSE TRANSCRIPTASE the family includes a number of significant pathogens, typically causing tumors or affecting the function of the immune system, e.g. HIV.

Question 24

What are inhibitors?

Answer 24

Chemical or biological molecules that regulate chemical reactions by slowing down or blocking them from occurring.

Question 25

What are the 3 types of Enzyme inhibition?

Answer 25

Competitive Inhibition:
- inhibitors bind to the active site of the enzyme (either ES or EI forms)

Uncompetitive Inhibition:
- inhibitors bind to a separate site but only to ES complex (ESI can form)

Mixed Inhibition:
- inhibitors may bind to enzyme or ES

ES: Enzyme Substrate Complex
EI: Enzyme Inhibitor Complex
ESI: Enzyme Substrate Inhibitor Complex

Question 26

What is a modulator? Compare and contrast positive and negative modulators.

Answer 26

Modulator: Ligands (substances) that can act on different parts of receptors and regulate enzyme activity

Positive modulators improve the binding affinity (increase reaction rate)

Negative modulators are uncompetitive inhibitors (slow down reaction rate)

Question 27

What is HIV protease?

Answer 27

An enzyme that is required to cleave viral peptide chains at specific locations. Essential to virus replication.

Question 28

How does rational drug design help us treat HIV?

Answer 28

COMPETITIVE or NONCOMPETITIVE INHIBITION: We can design drugs that mimic the protein so that HIV protease can’t perform its function

Question 29

Define allosteric enzymes. What happens to allosteric enzymes when modulators bind? How do allosteric interactions affect enzyme activity?

Answer 29

Enzymes changing shape (with the help of modulators) in order to be able to bind to the substrate better

Question 30

What is a Nucleophile?
What is a Electrophile?

Answer 30

Nucleophiles: donate electrons
Electrophiles: receive electrons

Question 31

What does induced fit mean?

What does cooperativity mean?

Answer 31

Induced Fit: The enzyme itself changes to fit the substrate better

Cooperativity: The binding of a ligand to a protein alters the affinity for subsequent binding of the same ligand

Question 32

What is Protein Modification?

Answer 32

Addition of a molecule onto a protein
- typically used to regulate activity of the protein
- most are reversible

Question 33

What is the most common type of regulatory modification?

Answer 33

Phosphorylation

Question 34

What is Phosphorylation? Why is it important?

Answer 34

Adding a phosphate group to an amino acid.
- Kinase (forward), phosphatase (reverse)
- ATP is a coenzyme
- Activates and deactivates many proteins
- Important in many cell signaling pathways (sending a signal from outside the cell across the plasma membrane).

Question 35

What is Acetylation? What effect to acetyl groups have on wound-up DNA?

Answer 35

ADDING CARBONS AND OXYGENS TO SOMETHING

A reaction that introduces an acetyl functional group into a chemical compound.
- (Deacetylation is the removal of an acetyl group)
- Regulates the activity of chromatin by affecting how strongly the histones bind DNA

• Acetyl groups help us see genes expressed
  because it UNWINDS DNA HISTONES!!

Question 36

What is Methylation?

Answer 36

Opposite of Acetylation: keeps DNA wound up tight.

Question 37

What is Ubiquitination?

Answer 37

FLAGS SOMETHING FOR GARBAGE TO THROW AWAY

Enzymatic post-translational modification in which a ubiquitin protein is attached to a substrate protein.
- Ubiquitination generally targets proteins for degradation but sometimes it just alters the function of a protein.