RECOMBINANT PROTEINS
Source: Hindu
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Context
- Researchers at the Indian Institute of Science (IISc), Bengaluru, have developed an innovative method for the mass production of recombinant proteins, presenting a safer and more efficient alternative to existing techniques.
- This advancement addresses the hazards associated with methanol usage in recombinant protein production, proposing the use of monosodium glutamate (MSG) instead.
Details
Traditional Method: Methanol-Induced Production
Recombinant Proteins:
- Recombinant proteins are produced by genetically modifying bacterial, viral, or mammalian cells.
- Applications:These proteins are crucial for various medical applications, including vaccine antigens, insulin, and monoclonal antibodies.
- Yeast (Pichia pastoris/Komagataella phaffii):The most widely used organism for this production.
Methanol-Induced Process:
- Promoter Activation:The yeast contains a unique promoter (AOX promoter) that is activated by methanol, which codes for the enzyme alcohol oxidase (AOX).
- Process:
- The gene coding for the desired protein is inserted next to the AOX promoter in the yeast genome.
- Yeast cells are grown using glycerol or glucose.
- Methanol is added to activate the AOX promoter, inducing protein production.
Challenges with Methanol:
- Safety Risks:Methanol is highly flammable and hazardous.
- Oxidative Stress:Methanol metabolism produces hydrogen peroxide, which can damage yeast cells and recombinant proteins.
Novel Method: MSG-Induced Production
Monosodium Glutamate (MSG) as an Alternative:
- Discovery:MSG, a USFDA-approved food additive, can activate a different promoter in the yeast genome.
- Promoter:This promoter codes for the enzyme phosphoenolpyruvate carboxykinase (PEPCK).
- Process:
- The MSG activates the PEPCK promoter, leading to protein production similar to that induced by methanol.
- Advantages:
- Safety:MSG is non-flammable and safer to handle than methanol.
- Efficiency:The process eliminates the oxidative stress caused by methanol.
Applications
- Biotech Industries:The novel expression system can be utilized for the mass production of valuable proteins.
- Products:Includes milk and egg proteins, baby food supplements, nutraceuticals, and therapeutic molecules.
- Scalability:The method has the potential for widespread adoption in industrial settings due to its safety and efficiency.
- Innovation:Represents a significant advancement in biotechnological processes, offering a more sustainable and safer approach to recombinant protein production.
About Recombinant Proteins
- Recombinant proteinsare proteins encoded by genes that have been cloned in a system that supports the expression of the gene and the translation of messenger RNA.
- This allows for the production of large quantities of a protein that is otherwise difficult to obtain.
- Recombinant protein technology is pivotal in biochemistry, molecular biology, and medicine.
Production of Recombinant Proteins
Gene Cloning:
- Isolation of Gene:The DNA sequence encoding the protein of interest is identified and isolated.
- Vector Selection:The gene is inserted into a plasmid or other suitable vector that will carry the gene into a host cell.
- Transformation:The vector is introduced into host cells (bacteria, yeast, insect, or mammalian cells).
Expression Systems:
- Prokaryotic Systems (e.g., E. coli):Simple, fast, and cost-effective. Suitable for non-glycosylated proteins.
- Eukaryotic Systems:
- Yeast (e.g., Saccharomyces cerevisiae):Capable of 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 producing complex proteins with post-translational modifications similar to human proteins.
Protein Purification:
- Cell Lysis:Release of protein from host cells.
- Chromatography Techniques:
- Affinity Chromatography:Uses a specific ligand to purify the target protein.
- Ion Exchange Chromatography:Separates proteins based on charge.
- Size Exclusion Chromatography:Separates proteins based on size.
Protein Characterization:
- SDS-PAGE:Determines the molecular weight of the protein.
- Western Blot:Confirms the presence of the protein using specific antibodies.
- Mass Spectrometry:Provides detailed protein mass and structural information.
- Activity Assays:Assess the functional activity of the protein.
Applications of Recombinant Proteins
Therapeutics:
- Insulin:Recombinant human insulin for diabetes management.
- Erythropoietin:Used to treat anemia.
- Monoclonal Antibodies:Targeted therapies for cancer and autoimmune diseases.
Vaccines:
- Hepatitis B Vaccine:Produced using yeast expression systems.
- HPV Vaccine:Utilizes recombinant technology to produce virus-like particles.
Diagnostics:
- Enzymes:Used in diagnostic assays (e.g., ELISA).
- Antigens:Used to detect the presence of antibodies in blood samples.
Advantages
- High Yield:Large quantities of protein can be produced.
- Purity:Proteins can be purified to a high degree.
- Consistency:Batch-to-batch consistency in protein production.
Challenges
- Post-translational Modifications:Prokaryotic systems cannot perform all modifications.
- Solubility Issues:Some recombinant proteins form insoluble aggregates.
- Cost:Production in eukaryotic systems can be expensive.
Sources:
PRACTICE QUESTION Q. Recombinant protein technology is a cornerstone of modern biotechnology with vast applications in medicine, research, and industry. Comment. (250 Words) |