Recombinant Human Proteins (CIE A Level Biology)

• DNA that has been altered by introducing nucleotides from another source is called recombinant DNA (rDNA)
• If the organism contains nucleotides from a different species it is called a transgenic organism
• Any organism that has introduced genetic material is a genetically modified organism (GMO)
• Recombinant DNA has been used to produce recombinant proteins (RP), thus recombinant proteins are manipulated forms of the original protein
• Recombinant proteins are generated using microorganisms such as bacteria, yeast, or animal cells in culture. They are used for research purposes and for treatments (eg. diabetes, cancer, infectious diseases, haemophilia)
• Most recombinant human proteins are produced using eukaryotic cells (eg. yeast, or animal cells in culture) rather than using prokaryotic cells, as these cells will carry out the post-translational modification (due to presence of Golgi Apparatus and / or enzymes) that is required to produce a suitable human protein
• The advantages of genetic engineering organisms to produce recombinant human proteins are:
  • More cost-effective to produce large volumes (i.e. there is an unlimited availability)
  • Simpler (with regards to using prokaryotic cells)
  • Faster to produce many proteins
  • Reliable supply available
  • The proteins are engineered to be identical to human proteins or have modifications that are beneficial
  • It can solve the issue for people who have moral or ethical or religious concerns against using cow or pork produced proteins

Insulin

• In 1982, insulin was the first recombinant human protein to be approved for use in diabetes treatment
• Bacteria plasmids are modified to include the human insulin gene
  • Restriction endonucleases are used to cut open plasmids and DNA ligase is used to splice the plasmid and human DNA together

• These recombinant plasmids are then inserted into Escherichia coli by transformation (bath of calcium ions and then heat or electric shock)
• Once the transgenic bacteria are identified (by the markers), they are isolated, purified and placed into fermenters that provide optimal conditions
• The transgenic bacteria multiply by binary fission, and express the human protein - insulin, which is eventually extracted and purified
• The advantages for scientists to use recombinant insulin are:
  • It is identical to human insulin, unless modified to have different properties (eg. act faster, which is useful for taking immediately after a meal or to act more slowly)
  • There is a reliable supply available to meet demand (no need to depend on availability of meat stock)
  • Fewer ethical, moral or religious concerns (proteins are not extracted from cows or pigs)
  • Fewer rejection problems or side effects or allergic reactions
  • Cheaper to produce in large volumes
  • That it is useful for people who have animal insulin tolerance

Factor VIII

• Factor VIII is a blood-clotting protein that haemophiliacs cannot produce
• Kidney and ovary hamster cells have been genetically modified to produce Factor VIII
• Once modified these recombinant cells are placed into a fermenter and cultured
• Due to the optimal conditions in the fermenter, the hamster cells constantly express Factor VIII which can then be extracted and purified, and used as an injectable treatment for haemophilia
• The advantages for scientists to use recombinant Factor VIII are:
  • Fewer ethical, moral or religious concerns (proteins are not extracted from human blood)
  • Less risk of transmitting infection (eg. HIV) or disease
  • Greater production rate

Adenosine deaminase

• Adenosine deaminase (ADA) is an enzyme used to treat the inherited condition called Adenosine Deaminase Deficiency
• ADA Deficiency is a common cause of Severe Combined Immunodeficiency (SCID)
• This is because the immune system is damaged
• The larva of the cabbage looper moth has been genetically modified (using a virus vector) to produce the enzyme adenosine deaminase so that it can be used as a treatment whilst the patients wait for gene therapy or when gene therapy is not possible
• The advantages for scientists to use recombinant adenosine deaminase are:
  • Fewer ethical, moral or religious concerns (proteins are not extracted from cows)
  • Less risk of transmitting infection or disease (from cows)
  • More reliable production of enzyme
  • Faster to produce many proteins