Alanine scanning

The Alaninscan is a method in molecular biology and protein biochemistry to identify the essential for the function, shape and stability of proteins or peptides molecular components. It is based on the systematic sequential replacement of each amino acid or, alternatively, more amino acids of a peptide, protein or protein part against the amino acid alanine. An exchange has a functionally or structurally essential amino acid of a protein with alanine usually a loss of protein function result.

Principle

The Alaninscan is based on the systematic, sequential replacement of each amino acid to alanine. This substitution has the consequence that all affected side chain atoms are removed to a large extent functionally neutral methyl group. Theoretically, even a complete removal of the side chain by replacing the amino acid glycine is conceivable, but it has such a substitution, thanks to the conformational flexibility of the peptide bond of glycine have a strong influence on the secondary structure of the protein. Is an essential function for the amino acid alanine replacement, a loss of the protein function can be observed usually. Similarly, a loss of protein function in exchange of an essential for the internal structure or the amino acid form occur.

Methodology

Classical methods of Alaninscans a protein based on the sequential replacement of each amino acid with the help of site-specific mutagenesis. This method is very costly, since each modified protein must be separately constructed, expressed and analyzed. By combining random mutagenesis with the display method such as phage display, the amount of work can be reduced. Also combinatorial approaches of Alaninscans are possible and facilitate the study of protein function by simultaneous exchange of several amino acids. A Alaninscan a small peptide is possible using chemical synthesis.