by Introduction to Recombinant Protein Technology
Recombinant protein technology involves modifying biological systems to generate novel proteins not normally produced by those systems. This allows scientists to study and make use of proteins important for human health, industry, research, and more. The technique relies on genetic engineering and cellular machinery to mass produce proteins of interest. Let’s explore how this groundbreaking technology works and the impacts it is having.
Isolation and Cloning of Genes
The first step involves isolating the gene that codes for the target protein from its original organism. The gene contains the blueprint for amino acid sequence of the protein. Scientists use techniques like polymerase chain reaction (PCR) to amplify the gene from DNA. They then insert the gene into a vector, which is usually a small, self-replicating piece of DNA like a plasmid. This is done using special restriction enzymes that cut the vector and insert at specific sites. The vector, now containing the foreign gene, is introduced into a host cell.
Expression of Cloned Genes in Host Cells
Bacteria are commonly used as host cells as they can be grown rapidly in culture. The most widely used bacterial host is Escherichia coli due to its fast growth rate, easy manipulation, and well-understood genetics. When the vector containing the cloned gene enters the E. coli cell, it directs the host’s cellular machinery to produce the target protein. RNA polymerase produces messenger RNA (mRNA) from the gene, which is then used as a template by ribosomes to synthesize the protein through amino acid assembly.
Isolation and Purification of Recombinant Proteins
To isolate the Recombinant Protein, the bacterial cells are lysed to break them open and release cell contents. The lysate containing the protein is then subjected to purification steps like centrifugation, chromatographic techniques, and electrophoresis to separate and concentrate the protein from other cellular proteins and compounds. Tags added during cloning help isolate the protein by binding to affinity columns. Highly purified recombinant protein is obtained through a series of purification protocols.
Applications and Benefits of Recombinant Protein Technology
Some key applications and benefits of recombinant protein technology include:
– Production of insulin and other hormones for medical treatments. Prior to recombinant methods, insulin was extracted from animals in limited quantities at high costs. Mass production lowered treatment costs.
– Production of enzymes used as industrial catalysts like chymosin in cheese production and lipase in detergents. Bacteria and yeast turn out enzymes much more efficiently than extracting from source organisms.
– Production of vaccines against diseases like hepatitis B. Recombinant subunit vaccines contain only the effective antibody-stimulating parts of a pathogen and are safer than whole pathogen vaccines.
– Production of proteins for scientific research applications like fluorescent tags and affinity ligands that have revolutionized techniques like microscopy, chromatography, and blotting assays.
– Potential production of insecticidal proteins from Bacillus thuringiensis for use as biopesticides and production of industrial enzymes for biofuels, bioplastics and more.
Challenges and Limitations
Some challenges include ensuring proper folding, structure and post-translational modifications of proteins produced in bacteria which lack certain eukaryotic processing abilities. Toxicity of highly expressed genes to host cells can also reduce yields. Scale-up to industrial levels further poses challenges in growing and handling large volumes of cultures. However, ongoing research aims to overcome such issues through strain and process engineering.
Future Prospects
As techniques advance, more complex eukaryotic proteins will become accessible through expression in mammalian or plant host systems. Production of biologics and replacement enzymes as industrial commodities holds promise. Gene therapy may leverage this technology to treat genetic disorders through direct mRNA or vectored protein delivery. Recombinant protein technology has already revolutionized biomanufacturing and will remain integral to translational life sciences going forward. With continued improvements, its full potential is yet to be realized.
*Note:
1. Source: Coherent Market Insights, Public sources, Desk research
2. We have leveraged AI tools to mine information and compile it
