Protein engineering is the foremost among the much recent advancement in biotechnology. It reflects some of the exciting possibilities that come out of deeper understanding of living systems.
Protein-engineering and design refer to the possibility of actually changing the structure of a protein at one’s desire so that it may function in a better way.
In a folded form the protein stability is a balance between the stabilising interactions (e.g. mainly hydrophobic) and the tendency towards destabilising (which is caused by the loss of conformational entropy as the protein adopts the unfolded form).
A reduction in entropy difference between folded and unfolded states would require a drastic reduction of the number of possible unfolded conformations. By substituting amino acids the stability of a protein can be changed.
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The substituted amino acids favour the stabilising interactions in a folded protein or destabilising interactions to some extent in a native protein. Many enzymes and proteins have been attempted to bring about changes so that improvement in their catalytic potency, thermal and pH stability, solvent tolerance, etc. could be made satisfactorily.
1. Differences between Protein Engineering and Protein Design :
Protein engineering differs from protein design. In protein engineering an existing protein is changed chemically or genetically so that its function can be changed as per desire. In protein design, the aim of the process is to fulfill a desired function.
Protein engineering is a multidisciplinary science. The engineering and design flow chart requires the interaction of physicists, biophysicists, molecular biologists, organic chemists and computer scientists.
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2. Steps of Protein Engineering :
The steps of protein-engineering are given below:
(i) Identification:
Protein -engineering starts with identification of a protein with a specific function which is to be changed accordingly to ones need.
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(ii) Isolation and Characterisation:
The identified proteins are isolated and characterised biochemically. Then 3D structure and function of protein and relationship between structure and function are found out.
(iii) Protein Modification:
Based on the above facts and known principles of protein conformation, modification in protein can be suggested which may bring about desired changes.
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(iv) Incorporation of Changes:
Changes are incorporated into the protein by side-directed mutagenesis or biochemical or molecular techniques (see preceding section). The activity of a new protein must be tested. Protein design starts from knowledge of principles of protein structure.
A sequence of amino acid is designed applying these principles with the objectives that it will take up a specific 3D structure and cause the protein for necessary function.
Appropriate DNA is synthesised as per amino acid of choice. Then it is cloned into expression vector system by recombinant DNA technology. The expressed protein will undergo tests and next stage of the design process.