Studying protein–protein interactions using peptide arrays

Abstract

Screening of arrays and libraries of compounds is well-established as a high-throughput method for detecting and analyzing interactions in both biological and chemical systems. Arrays and libraries can be composed from various types of molecules, ranging from small organic compounds to DNA, proteins and peptides. The applications of libraries for detecting and characterizing biological interactions are wide and diverse, including for example epitope mapping, carbohydrate arrays, enzyme binding and protein-protein interactions. Here, we will focus on the use of peptide arrays to study protein-protein interactions. Characterization of protein-protein interactions is crucial for understanding cell functionality. Using peptides, it is possible to map the precise binding sites in such complexes. Peptide array libraries usually contain partly overlapping peptides derived from the sequence of one protein from the complex of interest. The peptides are attached to a solid support using various techniques such as SPOT-synthesis and photolithography. Then, the array is incubated with the partner protein from the complex of interest. Finally, the detection of the protein-bound peptides is carried out by using immunodetection assays. Peptide array screening is semi-quantitative, and quantitative studies with selected peptides in solution are required to validate and complement the screening results. These studies can improve our fundamental understanding of cellular processes by characterizing amino acid patterns of protein-protein interactions, which may even develop into prediction algorithms. The binding peptides can then serve as a basis for the design of drugs that inhibit or activate the target protein-protein interactions. In the current review, we will introduce the recent work on this subject performed in our and in other laboratories. We will discuss the applications, advantages and disadvantages of using peptide arrays as a tool to study protein-protein interactions.

Modular peptides from the thermoplastic squid sucker ring teeth form amyloid-like cross-β supramolecular networks

“…A combinatorial peptide macro array interaction assay [10,[26][27][28][29] with a suckerin-peptide based library was designed in order to obtain a preliminary screening of peptide interactions and to identify binding interactions of binary peptide combinations ( Figure 1B, details of the assay are described in Materials and methods section). All binary combinations of peptides were screened at three different pHs and three analyte concentrations to generate 441 different conditions.…”

Selective Functionalization of Microstructured Surfaces by Laser‐Assisted Particle Transfer

“…[1] Recently, microstructures were also used to analyze single cells [2] or to screen the functional heterogeneity of enzyme activity on single molecule level, [3] as well as studying the impact of geometrical constraints and ligand specificity on stem cells. [4] Although the technology for combinatorial chemistry, based on the solid phase synthesis principle of Merrifield, [5] is well developed, especially for peptide arrays, [6] the functionalization of microcavities in the ultra-high density array format remains challenging.…”

Identifying Protein-protein Interaction Sites Using Peptide Arrays

“…Unlike other biochemical or biophysical methods for detecting and analyzing protein-protein interactions, peptide array screening requires a very low concentration of protein and can detect very weak binding. Peptide arrays can be used for many applications in peptide-protein interactions such as mapping of protein-protein or receptorligand interaction sites 4 , homo-or hetero-oligomerization interfaces, characterizing antibodies epitopes 5 , studying enzyme activities 6 and high throughput structure-activity relationship (SAR) studies 7 . For an in-depth review about peptide array screening see Katz et al 4 Several types of peptide arrays currently exist.…”

“…For an in-depth review about peptide array screening see Katz et al 4 Several types of peptide arrays currently exist. There are two major synthetic strategies for making peptide arrays: synthesis of the peptides before attaching them to the solid support, or synthesis of peptides directly on the solid support, mainly using the SPOT technique 4,8 . The peptides are synthesized on the solid support usually by 9-fluorenylmethyloxycarbonyl (Fmoc) chemistry 8 .…”

“…The peptides are synthesized on the solid support usually by 9-fluorenylmethyloxycarbonyl (Fmoc) chemistry 8 . Among the common synthetic schemes are peptide attachment through the N terminus (e.g., JPT pepstar arrays 9 ) and peptide attachment through the C terminus (e.g., PEPSCAN pepchip arrays 10 , JPT pepspot arrays 9 and INTAVIS celluspot arrays 11 ) 4 . The solid support can vary and so does the chemistry of the peptide coupling to it.…”