Unified schedule for multiple parallel solid-phase synthesis of so-called “difficult” peptides on polypropylene pins was developed. Increase in the efficiency of 9-fluorenyl(methoxycarbonyl) N-terminal amino-protecting group removal was shown to have a greater influence on the accuracy of the “difficult” peptide synthesis than the use of more efficient amino acid coupling reagents such as aminium salts. Hence the unified schedule for multiple parallel solid-phase synthesis of “difficult” peptides included the procedure for N-terminal amino group deprotection modified by applying a more efficient reagent for the deprotection and the standard procedure of amino acid coupling by carbodiimide method with an additional coupling using aminium salts, if necessary. Amino acid coupling with the help of carbodiimide allows to follow the completeness of the coupling via the bromophenol blue indication, thus providing the accuracy of the synthesis and preventing an overexpenditure of expensive reagents. About 100 biotinylated hepatitis C virus envelope protein fragments, most of which represented “difficult” peptides, were successfully obtained by synthesis on pins with the help of the developed unified schedule. 1. Introduction Development of proteomic and interactome research linked to the mass-spectral detection and amino acid analysis of peptide fragments of proteins requires extensive development of multiple solid-phase peptide synthesis in order to prepare huge sets of peptides used as calibration standards and as affinity ligands for interactome analysis and interaction site mapping [1–5]. These peptide sets are expected to contain up to several hundreds of peptides including those with modified side-chain functional groups, since the analysis of a single tissue sample from a single organism may require the preparation of more than a hundred of the so-called characteristic peptides (unique fragments of proteins under study). The field of peptide scanning usage, which includes multiple parallel peptide syntheses as an obligatory part of the method, also expands. Besides scanning proteins for B- and T-epitope motifs [6–12], kinase phosphorylation and other posttranslational modification sites [13–17], and studies of protease cleavage specificity [16, 18], multiple parallel peptide synthesis is employed for the search of antibacterial peptides [19], receptor peptide ligands [20], and preparation of novel biomaterials based on readily structured peptides and peptoids [21]. Though immunochemical research sometimes allowed the use of peptide preparations with
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