Lecture 21: PRODUCTION OF THERAPEUTIC PROTEINS WITH A NEW FUNCTION

Question 1

What influences the host choice for protein production?

Answer 1

Speed and cost (financial reasons) and glycosylation and folding (biochemical reasons)

Question 2

What is the worst to best host with relation to speed?

Answer 2

transgenic, plants, mammalian, insect, yeast, bacteria

Question 3

What is the worst to best host with relation to cost?

Answer 3

transgenic, mammalian, plants, insect, yeast and bacteria

Question 4

What is the worst to best host with relation to glycosylation?

Answer 4

bacteria, yeast, plants, insect, transgenic, mammalian

Question 5

What is the worst to best host with relation to folding?

Answer 5

bacteria, yeast, plants, insect, transgenic, mammalian

Question 6

What is the PTM for insulin?

Answer 6

Disulphide bonds

Question 7

What is insulin best produced in?

Answer 7

A and B chains in separate bacteria

Question 8

What is the PTM for EPO?

Answer 8

Glycosylation

Question 9

What is EPO best produced in?

Answer 9

Mammalian cell culture

Question 10

What is the PTM for antithrombin?

Answer 10

Gamma carboxylation

Question 11

What is antithrombin best produced in?

Answer 11

Transgenic animals

Question 12

How can we make new proteins?

Answer 12

Use a plasmid with promoter (cell type specific), transgender and antibiotic resistance for amplification in bacteria.

Question 13

What can we do to create mutations in proteins?

Answer 13

Amplify a known enzyme using degenerate PCR (with ineffective enzymes)

Question 14

What can be done with mutations with improved function?

Answer 14

They can be added to a plasmid using a promoter specific to bacterial or mammalian cels depending on whether or not post-translational modifications are required

Question 15

What do recombinant DNA technologies in the clinic require?

Answer 15

Repeated administration of drugs for the life of the patients/illness

Question 16

How can we provide a permanent treatment?

Answer 16

Use patients as the host for protein production

Question 17

What happens in gene therapy?

Answer 17

Plasmids are carried into patients using viral vectors as if pure plasmid entered, the body would degrade it.

Question 18

What happens with the gene in the viral vector?

Answer 18

It is inserted into the host cell chromosome

Question 19

What do viruses have?

Answer 19

Different types of tropisms to make them more effective to certain cells

Question 20

What does conventional biotechnology use?

Answer 20

A recombinant protein

Question 21

What does gene therapy use?

Answer 21

A gene that encodes the recombinant protein

Question 22

What is the problem with permanently treating type 1 diabetes?

Answer 22

Beta cells in the pancreas are destroyed so the autoantibodies recognise proteins on beta cell surface so no insulin is produced

Question 23

What is the solution to permanently treating type 1 diabetes?

Answer 23

Turn liver cells into insulin making machines

Question 24

What is the promoter for treating type 1 diabetes?

Answer 24

Glucose responsive (when glucose levels are high) and also liver cell specific

Question 25

What is the gene for treating type 1 diabetes?

Answer 25

pre-pro insulin cDNA

Question 26

What is the tropism for the virus for treating type 1 diabetes?

Answer 26

Liver cells