Chassis Effects

Question 1

Define a chassis

Answer 1

A structural framework of a system which is able to support other parts and devices

Question 2

Define a biological chassis:

Answer 2

a cell (bacterial, algal, fungal, mammalian) capable of living, replicating and expressing the synthetic biology parts/devices we introduce

Question 3

What is already present in the chassis?

Answer 3

Gives you the functions that make sure the cell can already survive, reproduce respire etc.

Question 4

Define metabolic burden

Answer 4

The amount of resources (raw materials and energy) taken from the host for maintaining and expressing foreign DNA and producing novel compounds

Question 5

Explain specific burden

Answer 5

Resources for the new function you have inserted are redirected from respiration (reduction of resources for directly linked metabolic pathways)

Question 6

Explain generic burden

Answer 6

the reduction in growth rate affects the rate of DNA and cell wall turnover indirectly (reduction of energy available for cell division)

Question 7

What is Artemisinin?

Answer 7

An anti-malarial drug

Question 8

What is the disadvantage of synthetic artemisinin production?

Answer 8

It is expensive

Question 9

What is the name of the plant that naturally produces artemesinin?

Answer 9

Sweet wormwood

Question 10

What is the problem with harvesting artemisinin from sweet wormwood?

Answer 10

The price will fluctuate with the weather and any major weather events due to the variation in crop growth

Question 11

What did Youyou Tu win a Nobel Prize for in 2015?

Answer 11

For showing that the active ingredient in the sweet wormwood extract was arteseminin

Question 12

Who was the first person to demonstrate that artemisinin could be made in ecoli?

Answer 12

Jay Keasling

Question 13

Which two compounds are the core precursers for producing artemisinin?

Answer 13

IPP and DMAPP

Question 14

What gene is ecoli missing to produce artemisinin?

Answer 14

ADS

Question 15

What is FPP used for in both sweet wormwood and ecoli?

Answer 15

Sweet wormwood - artemisinin

Ecoli - making cell wall components

Question 16

Which pathway produces FPP in ecoli?

Answer 16

The DXP pathway

Question 17

Which pathway produces FPP in yeast?

Answer 17

The mevalonate pathway

Question 18

What was the effect of putting all the genes that create FPP in yeast into ecoli?

Answer 18

The specific burden was too great and production slowed.

Question 19

What is the function of FPP in yeast?

Answer 19

Makes the cell wall

Question 20

Why does producing artemisinin in yeast have a lesser metabolic burden than in ecoli?

Answer 20

Because you are introducing less new proteins into the organism

Question 21

What other device was introduced into yeast to decrease the amount of organic synthesis required and how?

Answer 21

CYP by converting amorphadiene to a compound closer to artemisinin (artemisinic acid)

Question 22

What process was carried out to increase the specific yield of artemisinic acid production in yeast?

Answer 22

Decreased the expression of the enzymes that produced ergosterol by using a weaker promotor, so not as much as was usually used to sustain cell wall synthesis was produced.

Question 23

What process was carried out to remove the bottleneck in FPP production in yeast?

Answer 23

Increased the copy number of the enzyme to catalyse the bottleneck reaction was inserted so that twice as much existed and more mevalonate was produced and therefore more FPP.

Question 24

What is the compound that is the start block for artemisinin production?

Answer 24

Acetyl-CoA

Question 25

How was the production of Acetyl-CoA increased?

Answer 25

By using the technique of changing promoters and copy numbers

Question 26

What is a synthetic scaffold?

Answer 26

A protein (not an enzyme so it doesn’t have a function as such) that has pockets for the other proteins in the pathway to bind to, which increases their proximity. Increasing the proximity of the enzymes increases the reaction rate because the local concentration of the product will be quite high so the next step of the reaction will proceed faster.

Question 27

What does Jay Keasling have to say about modern synthetic biology methods?

Answer 27

That the artemisinin project would not have taken him 10 years had the tools such as abstraction, modularisation and standardisation had existed when he started.

Question 28

When can the synthetic biology way of making artemisinin be used?

Answer 28

Only when the natural stock of sweet wormwood is depleted and there is not enough artemisinin available