Protein Engineering

The process of developing proteins having desired functions by manipulating their stability and specificity is termed protein engineering. This can be done by taking two main approaches, rational design and irrational design (or directed evolution). In the former case, knowledge of the protein structure and function is considered and a rational gene mutation is planned.

Rational Protein Design
Knowledge based engineering of protein with desired characteristics

Design/Modelling (often Computer aided)
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Generate required changes in cloned DNA construct
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Express protein
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Purify protein
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Assess protein for desired changes

Irrational Protein Design
High throughput protein engineering

Generate library of DNA constructs (sometimes designed by Bioformatics)
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Express all constructs
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Screen protein with designed characteristics (hit) by high throughput
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Get DNA construct for hit for large scale expression

This is done by making rationally designed changes in the gene of the protein cloned in expression vector of heterologous expression. The production of protein molecules is changed by site directed or site-specific mutagenesis of their genes. While there are cases in which protein structure is not available, and thus irrational design is required in which random changes (mutation) are made in the protein and a mutant with desired properties is selected.

Objectives

Enzymes, antibodies, hormones, receptor proteins, etc., are the protein classes chosen for engineering. Out of these, enzymes are the most widely used ones. Protein engineering mainly aims to get functionally more useful and efficient proteins and obtain economic gains from them.

Protein engineering has the objectives of:

Increasing the stability of enzymes,
Increasing the catalytic efficiency of enzymes,
Introducing new active sites and new catalytic activity in enzymes,
Altering $pH$ dependence of enzymes,
Improving thermostability and modifying substrate specificity of enzymes,
Understanding structural and functional relationship for an enzyme for making predictions of actions,
Transferring the ability to make desirable enzyme in easy to cultivate and safer to use microbes,
Producing tailor-made enzymes in large quantities by increasing the expression of structural genes,
Producing hybrid enzymes,
Making isolation and purification of enzymes easy,
Making hormones resistant to attack by antibodies or stomach enzymes,
Getting more specific and potent biopharmaceuticals with altered pharmacological action, and
Getting humanised antibodies with less immunogenicity.