Protein Engineering: Different Biotechnology Applications

Dear Colleagues,

Protein Engineering is an emerging branch of engineering that applies knowledges of mathematics and molecular biology to the design of proteins. It operates iteratively, following a cyclic process that alternates stages in which the changes to be made are planned and executed with others in which the result of the changes is evaluated. There are two methods for the design of proteins: rational design and directed evolution. In the "rational design" method, changes are introduced in certain amino acids through directed mutagenesis, on the basis of the hypothesis that some specific changes will cause the desired functional effect. In some occasions, these changes occur when domains or structural motifs of different proteins are combined, generating a hybrid protein or chimera. This is a simple and cheap method. In the “directed evolution” method, random mutations are introduced in the protein under study, and only those variants that exhibit the desired properties are selected. Generally, two molecular biology techniques are used to perform random mutagenesis of isolated genes. One is known as "error-prone PCR" and consists in the amplification by PCR (polymerase chain reaction) of the gene that codes for the protein of interest in conditions that induce the DNA polymerase to make mistakes. The other procedure is called "DNA-shuffling" and consists in the fragmentation of the sequence to mutagenize by digestion with DNase, followed by a reassembly of the same sequence through PCR. Several rounds of mutation and selection give rise to a collection of modified proteins having the desired characteristics; however, some functional assays can be considerably complex, and the attainment of the mutant protein containing the desired modification can involve a very high number of assays. To solve these difficulties, the use of robotic processes or "high-throughput screening" is gaining momentum. These two techniques are inspired by natural evolution and sexual reproduction, respectively.The final or biochemical phase of the design cycle has the immediate objective of purifying the protein, as a preliminary stage for the resolution of its three-dimensional structure, since the determination of the structure of a protein is the best available tool to explain how it performs its function. The close link between the structure of a protein and its function makes solid structural information essential for the success of any approach to protein engineering. It is common for researchers to apply both techniques in the design of a given protein: first the rational design is applied, and then the product is subjected to directed evolution, following the design cycle. 

Dr. Benevides C. Pessela
Guest Editor

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