Biotechnology

Question 1

What is BIOLEACHING?

Answer 1

The process of oxidising insoluble metals ores (eg sulphides) by bacteria, into a soluble form (eg sulphate) which can be washed out of rocks using water.

Question 2

What characteristics does a bacteria need to be suitable for bioleaching? (2)

Answer 2

1) Can survive in highly acidic conditions, as sulfuric acid is produced as a result of the reaction
2) They need to be able to work in a fairly wide range of temperatures

Question 3

What are the advantages and problems of bioleaching? (5)

Answer 3

1) Gets metal from low grade ores in sufficient quantities to make a profit (not possible with traditional mining methods)
2) Can be used to extract valuable metals from waste/ash.
3) Does not produce sulfur dioxide (unlike conventional methods).
4) Bioleaching can be used in situ
5) Potential problem: production of sulfuric acid, which could seriously harm living organisms.

Question 4

Describe the method of batch cultures, and the difference between standard and fed batch cultures

Answer 4

Microorganisms can be grown in fermenters. In batch cultures, when the maximum amount of product is produced, the fermentation is stopped and the product harvested. The fermenter is cleaned out, and then a new culture is started.

In standard batch culture, the fermentation is left to proceed, nothing is added our taken out, Waste products are allowed to escape.

In fed batch culture, a carbohydrate source is added (eq corn syrup), the fermentation is kept going for longer, and therefore more economic.

Question 5

What are the requirements for ANY type of fermentation? (5)

Answer 5

1) Carbohydrate (energy source)
2) Nitrogen (protein synthesis)
3) Air (for aerobic bacteria)
4) Cold sleeves of water (reactions generally exo)
5) Use of pH buffers

Question 6

What is characteristic of a CONTINUOUS batch culture?

Answer 6

There is NO down time between batches. Small vessels are used to give yield as product is drawn off.

Question 7

What are the advantages/disadvantages of a batch culture?

Answer 7

1) Fermenter can be used for different cultures
2) If contamination occurs, only one batch is lost
3) Few Technical issues, easily set up

4) Downtime between setups

Question 8

What are the advantages/disadvantages of a continuous culture?

Answer 8

1) No ‘down time’
2) Relatively small vessels can be used, less space is needed to grow enough microOrg to give a good yield.

3) If contamination occurs, many more batches are affected.
4) Cells sometimes clump together and block inlet/outlet pipes.

Question 9

What is the fungus used to manufacture penicillin?

Answer 9

Penicillium

Question 10

Why is penicillium a secondary metabolite?

Answer 10

Penicillin production only begins after the fungus has been growing for a while.

Question 11

How is penicillin produced?

Answer 11

The Penicillium fungus is grown in a FED BATCH CULTURE until the maximum amount of penicillin has been produced. The fermentation is stopped and the antibiotic harvested, fermenter is cleaned out and a new culture is put in. Process starts again.

Question 12

How are enzyme producing bacteria grown?

Answer 12

They are usually grown in BATCH CULTURE.

They are provided with a carbon source (waste product of industrial process eg remains of sugar cane, or meal of soya beans) and a nitrogen source (protein, like urea, or remains of yeast cells).

Bacteria are aerobic, contents of fermenter are well aerated.

Question 13

How are enzymes extracted? (3)

Answer 13

Some organisms secrete the enzymes into medium around them, but in some cases, enzymes remain inside the cells.

After fermentation has finished, the culturei s heated to kill the cells. If enzymes are in cells, they are broken open, allowing enzymes to escape from cells and dissolve in culture medium. This is concentrated and filtered. Cell fragments left behind. Enzyme collected in solution.

Enzymes can be purified (if required) and packaged in a easily usable form.

Question 14

What organism is MYCOPROTEIN?

Answer 14

A fungus called Fusarium, made up of long thin threads of HYPHAE

Question 15

How is mycoprotein grown?

Answer 15

It is grown in a CONTINUOUS BATCH CULTURE. There is a steady input of nutrients (glucose for respiration, carbon, protein ammonium phosphate and fat molecules) and steady harvest of fungus. Temperature. pH and oxygen content are kept constant.

Unlike penicillin, no stirrer is used.

Question 16

How is mycoprotein processed?

Answer 16

Liquid culture containing fungi run off from base of fermenter and centrifuged to separate. Enzymes are used to break RNA of the hyphae down. Filtration and steam treatment complete the process.

Question 17

What is a BROAD SPECTRUM ANTIBIOTIC?

Answer 17

An antibiotic that acts against a wide range of disease-causing bacteria

Question 18

What are bacterical cell walls made of?

Answer 18

Peptidoglycans

Question 19

What is the function of glycoprotein peptidases?

Answer 19

They are an enzyme which builds crosslinks that hold together peptidoglycans in a bacterical cell wall.

Question 20

What are autolysins?

Answer 20

An enzyme secreted by growing bacteria. It makes little holes in its cell wall, allowing the wall to stretch. New peptidoglycan chains link up across it.

Question 21

What is the mode of action of penicillin?

Answer 21

It inhibits glycoprotein peptidases, preventing peptidoglycan chains linking up. Autolysins keep making new holes, so cell wall becomes weaker. Eventually, the cell bursts due to osmosis of water into the cell.

Question 22

What organism does penicillin affect, and why does it no affect other types of cells?

Answer 22

Human cells don’t have cell walls, and viruses do not have cells.

Question 23

What causes antibiotic resistance?

Answer 23

In a colony of several million bacteria, one or two may have RESISTANCE GENES in their cells. This gene produces enzyme BETA-LACTAMASE, which breaks up penicillin molecules.

Another example is in the bacterium Staphylococcus. Some produce enzyme penicillinase (inactivates penicillin).

Question 24

What spreads antibiotic resistance? (2)

Answer 24

1) Resistance genes are usually carried on plasmids. When the few resistant bacteria with mutant genes survive antibiotic exposure, they are free to breed, producing a whole population that have inherited the resistance gene.
2) Plasmids can be transferred between different species of bacteria, this increases chances of resistant strains developing.
3) Unrelated bacteria can gain resistance from their neighbours through HORIZONTAL GENE TRANSFER.

Question 25

How can antibiotic resistance be prevented? (2)

Answer 25

1) Finishing your entire course of antibiotics

2) Doctors keep one or two antibiotics in reserve, only using them when every other antibiotic has failed.

Question 26

How are enzymes immobilised?

Answer 26

Enzyme is mixed with sodium alginate, and dripped into calcium chloride.

Question 27

What are the advantages of immobilised enzymes?

Answer 27

1) Easy to purify the product, compared to mixing the enzyme with its substrate.
2) Immobilised enzymes are more tolerant to temp and pH changes. (held firmly in shape by alginate, and not fully exposed to temp of pH changes)
3) Enzymes are easily recovered, more economical

Question 28

How does GLUCOSE OXIDASE (or peroxidase enzymes) work, and how is it used?

Answer 28

Glucose oxidase is immobilised and stuck to a pad on the surface of a dipstick. it oxidises glucose into gluoconolactone, and produces hydrogen peroxide. A colourless chemical on the pad detects hydrogen peroxide and changes colour to brown (the more glucose, the darker the colour).

Question 29

How do biosensors work?

Answer 29

Also relies on glucose oxidase. The reaction produces a tiny electric current, picked up by an electrode on the test strip. The more glucose the greater the current.

Question 30

What are problems with producing MONOCLONAL ANTIBODIES on a large scale?

Answer 30

B lymphocytes (manufactures antibodies) which divide, do not produce antibodies. and B lymphocytes which produce antibodies do not divide.

Question 31

What are HYBRIDOMA cells?

Answer 31

Plasma cells fused with cancer cells which divide indefinitely and secrete antibodies.

Question 32

How are hybridoma cells produced? (4)

Answer 32

1) Antigen injected into mouse
2) Mouse B lymphocytes recognise antigen proliferate and form plasma cells.
3) Plasma cells are taken from spleen of mouse and fused with cancer cells.
4) Tiny samples taken, only one cell present in each well. Every well tested so that any antibody producing hybridoma cells can be found.

Question 33

What is hCG?

Answer 33

Human Chorionic Gonadotrophin, a hormone released during pregnancy.

Question 34

How are antibodies specific to hCG produced? (3)

Answer 34

1) hCG injected into mouse
2) an antibody producing leucocyte is fused with a mouse myeloma cell
3) Cell is grown, forms a clone in which all cells secrete monoclonal antibodies

Question 35

How does a pregnancy testing dipstick work?

Answer 35

1) hCG specific antibodies bound to particles of gold, antibody-gold complexes coat the end of the dipstick
2) A second monoclonal antibody which specifically binds with hCG-antibody-gold is immobilised and coats a region further up (patient test region)
3) Dipstick is dipped into urine, any hCG binds with the first antibodies, and carried upwards.
4) hCG-antibody-gold complexes bind to second antibodies (immobilised). As more and more particles arrive, a pink colour (or other colour) builds up.
5) Control region contains third type of monoclonal antibody, immobilised even further up the stick. These antibodies are anti-mouse (from goats) and bind to antiody-gold complexes even with no hCG. Goes pink even of test is negative.

Question 36

How are monoclonal antibodies used in diagnosis? (3)

Answer 36

1) Used to locate blood clots (in person thought ot have deep vein thrombosis), Mouse injected with fibrin (main protein in blood clots). Antibodies produced labelled using radioactive chemical that produces gamma radiation.

Labelled antibodies introduced into patient’s blood, carried in bloodstream, and bind to any fibrin molecules. Gamma-ray camera then used to detect position of antibodies (therefore blood clots).

2) Can also be used to track down cancer cells (different proteins on cell surface memb from normal body cells)
3) Can determine exactly what strain of virus/bacterium is causing infection, and speeds up choice of most appropriate treatment.

Question 37

What are magic bullets, and how do they work?

Answer 37

Monoclonals used to deliver drugs. Once monoclonals have been made to match up protein found on surface of cancer cells, molecules of anti-cancer drugs are attached. Monoclonals injected into patient’s body, and attach only to surface of cancer cells. Drug destroys cancer cells but not other cells.

Question 38

What limits the success of ‘magic bullets’? (2)

Answer 38

1) Difficult to produce antibodies that bind only to cancer cells.
2) Monoclonals (from mice) are recognised as foreign by patient’s body, and destroyed before they can reach their target.