16 - Protein Engineering

Question 1

protein engineering

Answer 1

• used to make natural tools more useful tools
• rational methods
• non-rational methods

Question 2

subtilisin

Answer 2

• protease that removes protein-based stains by chopping up the protein

Question 3

protein design

Answer 3

• can be used to improve the stability of an enzyme

- changes an amino acid close to a catalytic region

Question 4

GFP revolution

Answer 4

allows biological processes to be highlighted

Question 5

GFP mutation

Answer 5

• highly mutated through rational engineering
• changing bases to His, Trp, Phe, or Tyr will change the luminescence colour of the protein
• mutating Ala to a larger, charged protein will repel dimers and produce a monomer

Question 6

rational protein engineering

Answer 6

using science based knowledge to alter proteins to be more useful

Question 7

non-rational protein engineering

Answer 7

• libraries of variants mutated and the successful variants are replicated
• empirical strategy where the solution is not known before-hand
• directed evolution

Question 8

human therapeutic antibodies

Answer 8

• therapeutic antibodies must be human antibodies that bind to human protein
• human immune system is unable to make antibodies against human proteins

1. microbial surface display
2. transgenic mouse
3. human memory B-cell immortalization
4. humanization

Question 9

microbial surface display

Answer 9

• libraries of antibodies displayed on the surface are examined
• links function with gene, therefore human antibodies can be created from it

Question 10

transgenic mouse

Answer 10

• mice immune system consists of human antibodies after human gene injection
• human antibodies are made from the mice antibodies

Question 11

human memory B-cell immortalization

Answer 11

isolate memory cells of an infected patient that survived, and make antibodies for another

Question 12

humanization

Answer 12

mice binding/recognition site are inserted into human genes

Question 13

avastin

Answer 13

• antibody used against tumors

- binds to vascular endothelial growth factor (VEGF) to prevent it from binding to the receptor

Question 14

vascular endothelial growth factor (VEGF)

Answer 14

• released by tumors

- stimulates angiogenesis

Question 15

angiogenesis

Answer 15

formation of new blood cells

Question 16

phage display

Answer 16

displaying foreign peptide on the surface of a filamentous phage

Question 17

filamentous phage

Answer 17

tubes with DNA that encode protein for structure

Question 18

phage display mechanism

Answer 18

1. insert genome into phage
2. transform prophage vectors to E. coli
3. phages will express different antibodies

Question 19

prophage

Answer 19

virus found in its DNA form, therefore it’s not infectious yet

Question 20

in vitro affinity selections

Answer 20

• phage display done 3-5 times on successful variants
• repeated for amplification
• DNA sequenced is used to find the actual sequence of the amplified structure

Question 21

Humira

Answer 21

• drug developed from phage display
• # 1 best selling drug
• binds to TNF-alpha to prevent it from binding with its receptor
• used to treat rheumatoid arthritis

Question 22

rheumatoid arthritis

Answer 22

immune system attacks cartilage and breaks it down

Question 23

infliximab

Answer 23

• synthesized through humanization
• works similarly to Humira (adalimumab)
• has more side effects
• decreased in sales