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Topic : Recombinant Protein Production

GS-3 Mains  : Science and Technology

Revision Notes

What are recombinant proteins?

  • Proteins made using genetically modified organisms (bacteria, yeast, or mammalian cells).
  • Used in various medical applications like vaccines, insulin, and monoclonal antibodies.
  • Produced by inserting the gene coding for the desired protein into a host cell.

Why is yeast (Pichia pastoris) preferred?

  • Most widely used organism for recombinant protein production.

Traditional method (methanol-induced):

  • Uses a promoter activated by methanol to induce protein production.
  • Challenges:
    • Safety hazard: Methanol is highly flammable and toxic.
    • Oxidative stress: Byproduct of methanol metabolism damages cells and proteins.

Novel method (MSG-induced):

  • Uses monosodium glutamate (MSG), a safe food additive, to activate a different promoter.
  • Advantages:
    • Safer: MSG is non-flammable and easier to handle.
    • More efficient: Eliminates oxidative stress from methanol.

Applications of the new method:

  • Mass production of valuable proteins in biotech industries.
  • Products: Milk and egg proteins, baby food supplements, nutraceuticals, and therapeutic molecules.
  • Scalable for widespread adoption due to safety and efficiency.
  • Significant advancement in biotechnology for a more sustainable and safer approach.

Recombinant protein production process:

  1. Gene Cloning:
    • Isolate the gene coding for the protein of interest.
    • Insert the gene into a vector (plasmid) for transfer into host cells.
    • Introduce the vector into host cells (bacteria, yeast, insect, or mammalian).
  2. Expression Systems:
    • Prokaryotic (e.g., E. coli): Simple, fast, and cost-effective for non-glycosylated proteins.
    • Eukaryotic:
      • Yeast (e.g., Saccharomyces cerevisiae): Can perform some post-translational modifications.
      • Insect Cells (e.g., Baculovirus system): High yield and proper folding for complex proteins.
      • Mammalian Cells (e.g., CHO cells): Best for complex proteins with human-like modifications.
  3. Protein Purification:
    • Release the protein from host cells through cell lysis.
    • Purify the target protein using chromatography techniques:
      • Affinity chromatography (specific ligand)
      • Ion exchange chromatography (protein charge)
      • Size exclusion chromatography (protein size)
  4. Protein Characterization:
    • SDS-PAGE: Determines protein molecular weight.
    • Western Blot: Confirms protein presence using specific antibodies.
    • Mass Spectrometry: Provides detailed protein mass and structure information.
    • Activity Assays: Assesses the functional activity of the protein.

Applications of recombinant proteins:

  • Therapeutics: Insulin, erythropoietin, monoclonal antibodies.
  • Vaccines: Hepatitis B, HPV.
  • Diagnostics: Enzymes and antigens used in assays.

Advantages of recombinant protein production:

  • High yield of proteins.
  • Highly purified proteins.
  • Consistent production across batches.


  • Not all post-translational modifications can be done in prokaryotic systems.
  • Solubility issues: Some proteins form insoluble aggregates.
  • Cost: Eukaryotic systems can be expensive.




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