A Novel Combinatorial Approach to High-Density Peptide Arrays

Abstract

Combinatorial synthesis of peptides on solid supports (1), either as spots on cellulose membranes (2) or with split-pool-libraries on polymer beads (3), substantially forwarded research in the field of peptide-protein interactions. Admittedly, these concepts have specific limitations, on one hand the number of synthesizable peptide sequences per area, on the other hand elaborate decoding/encoding strategies, false-positive results and sequence limitations. We recently established a method to produce high-density peptide arrays on microelectronic chips (4). Solid amino acid microparticles were charged by friction and transferred to defined pixel electrodes onto the chip's surface, where they couple to a functional polymer coating simply upon melting (Fig. 16.1 A-D,F). By applying standard Fmoc chemistry according to Merrifield, peptide array densities of up to 40,000 spots per square centimetre were achieved (Fig. 16.1G). The term "Merrifield synthesis" describes the consecutive linear coupling and deprotecting of L-amino acids modified with base-labile fluorenylmethoxy (Fmoc) groups at the N-terminus and different acid-sensitive protecting groups at their side chains. Removing side chain protecting groups takes place only once at the very end of each synthesis and generates the natural peptide sequence thereby.

Purification of Peptides in High-Complexity Arrays

“…[58] The established protocol for the silanization with the bromine silane was conceived for a 10 mM solution in anhydrous dichloromethane. [57][58]72] Compared to earlier protocols, the silane concentration had already been reduced by three quarters. [128] However, the synthesis of the bromine silane is labor-intensive and comparatively high amounts were needed to silanize the 22 x 21 cm² laser printer substrates.…”

“…[46,[56][57][58][59][60] In general, amino acid micro particles are triboelectrically charged in custom-built aerosol chambers and, then, transported to the surface of a semiconductor chip. The chip surface is equipped with a defined grid of aluminum pixel electrodes which can be selectively energized.…”

“…[45] In contrast, the CMOS micro chips are equipped with aluminum electrodes (Al/Al2O3, "Peptide Chip 5"). [57,59] …”

Genome-Wide Profiling of Molecular Recognition of Histone PTMs

“…Recent progress in array technology enables now generation of high-density arrays that may contain up to 40,000 peptides per cm 2 of solid support. Thus, such highdensity arrays would in principle enable a genome-wide analysis of reader domains with all so far identified potential peptidic recognition motifs (Beyer et al 2009). Current technology allows for parallel synthesis of peptides directly on a cellulose membrane.…”

Combinatorial/Library Peptide Synthesis

“…After coupling completion, the glass slide substrate is transferred to the compartment in which washing and deprotection is performed. By applying standard Fmoc chemistry, peptide array densities of up to 40 000 spots/cm 2 were prepared [185]. The scheme of the synthesizer is illustrated in Figure 12.5.…”