[15] Preparation of equimolar mixtures of peptides by adjustment of activated amino acid concentrations

The preparation and the use of defined mixtures of combinatorial libraries of peptides have been the areas of intense interest. In essence, there are two general methods for the preparation of such libraries. The first involves the direct coupling of a mixture of carboxy-activated amino acids, with the amino terminus (or termini) of a peptidyl solid support; the length of the mixture is increased by multiple such couplings. This method is extremely simple and can be performed in a standard peptide synthesizer, but is at some disadvantage in attempting to achieve equimolarity of the component peptides. The second method, often referred to as the “split resin method,” involves the coupling of multiple individual activated peptides with aliquots of peptidyl resin, followed by mixing the aliquots to form the mixture; repartioning of the mixed peptidyl resin and repetition of these steps provide the desired full-length mixture. This method has the distinct advantage over the former in that an equimolar mixture of peptides can be obtained, with the accuracy, in which the aliquots can be measured and coupling can be taken to completion. However, the split resin method is quite tedious, since each mixed position requires a number of individual coupling reactions equal to the number of components at that position. This chapter discusses the procedures for the preparation of peptide mixtures, using mixtures of activated amino acids. Although the procedures described here are for relatively simple mixtures, using specific blocking groups and carboxy-activating agents, diverse activating agents and blocking groups can be used; very large peptide mixtures have been made by the method.