Chapter 4

Question 1

Describe the structure of antibodies

Answer 1

made of two identical light chains and two identical heavy chains (longer, attached to carbohydrates) and have two antigen-binding sites with hypervariable loops

Question 2

What holds antibodies together?

Answer 2

disulfide bonds connect their beta sheets

Question 3

Define polyclonal as it relates to antibodies

Answer 3

many similar antibodies

Question 4

Define monoclonal as it relates to antibodies

Answer 4

many identical antibodies

Question 5

What type of cell is responsible for producing antibodies?

Answer 5

immune B cells

Question 6

Describe catalytic antibodies

Answer 6

bind to transition states, stabilizing them

Question 7

Describe how polyclonal antibodies are created

Answer 7

injecting an animal with an antigen over a period of weeks will cause polyclonal antibodies to secreted into the bloodstream

Question 8

Describe how monoclonal antibodies are created

Answer 8

fusing a B cell with a tumor cell will create hybrid cells that make monoclonal antibodies and divide forever

Question 9

Define immunoprecipitation

Answer 9

antibodies are added to a mixture of molecules, then collected with their antigen by centrifugation

Question 10

Define immunoaffinity column chromatography

Answer 10

This is a method for isolating proteins by filling a column with beads coated in antibodies. A mixture of molecules are poured in, then a wash to get rid of any stuck proteins, then the protein of interest is eluted.

Question 11

Define Western Blotting

Answer 11

This is a type of gel electrophoresis in which proteins move downward a vertical gel, with their speed determined by size and net charge.

Question 12

Describe the SDS Page gel

Answer 12

This is a type of gel electrophoresis in which proteins are first coated with SDS, then treated with Mercaptoethanol to break disulfide bonds and equalize the charge of each protein, so they move down the gel at a speed determined only by size.

Question 13

Describe how proteins can be identified in gels via antibodies

Answer 13

After a gel, proteins can be transported to a membrane that is incubated with antibodies that will bind to the protein of interest, revealing its position. Frequently, two antibiotics are used, one to bind to the protein, and another to bind to the first antibiotic.

Question 14

Describe how antibodies are used in microscopy

Answer 14

Fluorescent or gold-labeled antibodies can be combined with fluorescence or immunoelectron microscopy to detect proteins

Question 15

How do enzymatic proteins bind to their ligands?

Answer 15

Proteins bind to their ligands (ligand=any molecule bound to a protein) by electrostatic and hydrogen bonds (and other noncovalent).

Question 16

What are the three ways an enzyme can lower activation energy? What kind of reactions (delta G) do they catalyze?

Answer 16

binding two molecules together, rearranging electrons, or forcing substrates (substrates=ligands that bind to enzymes) into their transition states

do not always require ATP, as they also catalyze spontaneous reactions.

Question 17

Describe the binding sites of enzymes

Answer 17

an active site, where their substrate binds, and an allosteric site, where regulators bind

Question 18

Describe feedback inhibition

Answer 18

a type of negative regulation that occurs when a substrate binds to the allosteric site of an enzyme that was involved in its formation

Question 19

Describe positive regulation

Answer 19

Positive regulation occurs when an enzyme is stimulated by a regulatory molecule. This can be used to activate another branch in a synthesis pathway. This can also occur with the addition of a nucleotide such as ADP.

Question 20

Describe molecular switches

Answer 20

Molecular switches are intracellular signaling proteins that toggle between an active and inactive site in response to receiving a signal.

Question 21

Describe the enzyme lysozyme. Include: ligand, bond, basic type, and details of the reaction

Answer 21

Lysozyme changes the shape of the oligosaccharide to a strained conformation state and brings the glycosidic bond close to two of its amino acids.

One forms a transient covalent bond with the substrate and other becomes charged by donating an H and polarizes the water.

This allows a hydrolysis reaction to take place, which binds an oxygen-hydrogen to the substrate. This reaction also lets the acimo acids return to their original state.

Question 22

Describe competitive inhibitors and how they change Vmax or Km

Answer 22

Competitive inhibitors do not change Vmax, they just increase Km. The reaction needs more substrate to be as fast as it was before, but competitive inhibitors can be overcome with extra substrate.

Question 23

Describe noncompetitive inhibitors and how they change Vmax or Km

Answer 23

Noncompetitive inhibitors do not affect Km, but they do lower Vmax. Noncompetitive inhibitors function by removing the number of active enzymes. They disable enzymes by causing a conformational change.

Question 24

Define protein phosphorylation. Include purpose and catalysts.

Answer 24

Protein phosphorylation is the covalent addition of a phosphate group to a side chain of a protein. This can be done by removing a phosphate group from ATP.

Catalyzed by protein kinase and removed by protein phosphatases.

Phosphorylation can both stimulate or inhibit proteins, and be added or removed quickly.

Question 25

Define protein family

Answer 25

a group of polypeptides that shares a similar amino acid sequence or 3D structure, reflecting a common evolutionary origin

Individual members often have related but distinct functions, such as kinases that phosphorylate different target proteins

Question 26

Describe affinity chromatography and its use of isolating proteins that interact with the protein of interest

Answer 26

Affinity chromatography is used to separate polypeptides based on their ability to bind to a particular molecule. One example would be in immunoaffinity column chromatography. This can be used to isolate proteins that physically interact with the protein being studied by adding a mixture of proteins after the main protein has bound.

Question 27

Define cell extract

Answer 27

Cell extracts (or homogenates) are the first step in protein purification, where cells are lysed

Question 28

Define centrifugation

Answer 28

the most widely used procedure to separate a homogenate into different parts, or fractions

the homogenate is placed in test tubes and rotated at high speed in a centrifuge or ultracentrifuge

Question 29

Define chromatography (general)

Answer 29

a technique used to separate the individual molecules in a complex mixture on the basis of their size, charge, or their ability to bind to a particular chemical group

In a common form of the technique, the mixture is run through a column filled with a material that either binds or lets through the desired molecule

Question 30

Define electrophoresis (general)

Answer 30

a technique for separating a mixture of proteins or DNA fragments by placing them on a polymer gel and subjecting them to an electric field.

molecules migrate through the gel at different speeds depending on their size and charge