The structural basis of allergenicity: recombinant DNA‐based strategies for the study of allergens

Abstract

The use of recombinant DNA techniques for the study of allergenicity of proteins is a viable, and in many ways a preferred, alternative to the traditional procedures of protein purification, digestion and analysis of peptides for both allergenicity and amino acid sequence. The process of protein purification can be difficult and in many instances workers are forced to use only partially pure fractions that make the identification of the allergenic proteins uncertain. Furthermore, the purification and sequencing of peptides and their testing for retention of allergenic properties, represents a substantial and time-consuming work load. The synthesis of families of synthetic peptides to characterize the amino acids important for allergenic properties is also expensive and time-consuming. On the other hand, the preparation of a cDNA library from an allergen source is today a relatively easy and inexpensive task. The isolation and purification of cDNA clones is comparatively trivial compared to protein purification. Using the techniques described in this text, it can be seen that the molecular biological approach, although in some respects similar in principle to those of the protein chemist to study allergens, provides the capability to study several clones at the same time, and to compare clones for the presence of conserved regions corresponding to allergenic determinants. In addition, the techniques for generating mutant sequences provides perhaps the most powerful and simple set of procedures available for defining the amino acid structures essential for proteins or peptides to behave as allergens.