Biochemistry II lectures 1&2

Question 1

What are monoclonal antibodies?

Answer 1

-used as therapeutic agents
-produced from a single B-lymphocyte clone and bind the same epitope
-100 FDA approved mabs

Question 2

What are IgG antibody chains made up of?

Answer 2

• 1 light and 1 heavy chain (2 of each in total)
  -constant and variable sequences
  -Light Chain: VLCL
  Heavy Chain: VH3CH
  -disulfide bonds stabilize regions

Question 3

What are the fragments of Immunoglobulin?

Answer 3

-Fab (antigen recognition binding site)
-Fc (complement; activates complement-mediated cytotoxicity)

Question 4

How are antibodies flexible?

Answer 4

-loops between Ig domains (hypervariable)
-protease sensitive; cleaves peptides

Question 5

describe Fab (fragment antigen binding)

Answer 5

-fragment has 4 Ig domains
-2 Ig domains in each of the 2 polypeptide chains (1 Heavy; 1 Light chain)
-produced by proteolysis of whole antibody

Question 6

describe scFv (single-chain variable fragment)

Answer 6

-engineered
-2 variable domains (1 heavy; 1 light chain)
-domains fused together with linker region (10-25 amino acids)

Question 7

describe a Bispecific monoclonal antibody

Answer 7

-engineered
-2 antigen binding regions that recognize and bind 2 different antigens
-ex: BiTE; fusion of 2 different scFV’s and is bi-specific with one antigen binding region selective for T-Cells

Question 8

Describe a Trifunctional antibody

Answer 8

-engineered mab; 3 binding sites
-antigen binding site in variable domains recognize epitope (T Cell CD3; antigen from a tumor cell; Fc part of the mab designed to bind to normal Fc receptor)

Question 9

describe molecular recognition

Answer 9

-interaction of a protein molecule with a ligand (forms stable complex)
-measured by binding affinity
-high affinity=complex is favored over free P and L
-P and L binding is due to noncovalent interactions
-induced-fit structural changes

Question 10

What is Kd?

Answer 10

-equilibrium dissociation constant
Kd= [P][L]/[PL]
-smaller Kd value, higher affinity

Question 11

What is Specificity?

Answer 11

-affinity for one ligand over other ligands
-can also be affinity of ligand for one protein over another

Question 12

What does a low Km mean?

Answer 12

-high affinity for substrate binding

Question 13

Which choice best describes competitive binding between the enzyme substrate, S, and inhibitor, I?

Answer 13

-I binds the active site
-the region where I binds partially overlaps the active site
I induces a conformational change that prevents S binding

Question 14

What happens to the apparent Vmax and Km with no inhibitor?

Answer 14

Km and Vmax are the same

Question 15

What happens to the apparent Vmax and Km with competitive inhibition? (inhibitor binds only free E)

Answer 15

-Km is changed (Km[1+[I]/Ki)
-Vmax is same

Question 16

What happens to the apparent Vmax and Km with non-competitive inhibition? (inhibitor binds free E and ES complex with equal affinity)

Answer 16

-Km is unchanged
-Vmax is changed (Vmax/(1+[I]/Ki)

Question 17

What happens to the apparent Vmax and Km with uncompetitive inhibition? (inhibitor only binds to ES complex)

Answer 17

-Km is changed (Km/(1+[I]/Ki)
-Vmax is changed (Vmax/1+[I]/Ki)
-both apparent Vmax and Km change by the same amount

Question 18

What does a Ki value indicate?

Answer 18

-dissociation constant that describes the binding affinity between the inhibitor and the enzyme
-concentration of inhibitor required to produce half maximum inhibition