Target proteins selection

Disadvantages of both phage and mRNA display are the requirement for numerous rounds of selection and amplification, as well as the need to express and purify target proteins. Selectively infective phage (J3) and bacterial (H4, J2) and yeast (Y2) two-hybrid methods can overcome these obstacles because they are one-step screening assays with in vivo expressed target proteins. 

When 14-3-3s were first identified as phosphorylation dependent binding proteins (note that a selection of non-phosphorylated targets are known), target protein phosphorylation sites were mapped and it was immediately apparent that 14-3-3s bound preferentially to specific phosphorylation motifs. The advent of oriented... 

Although a number of cell lines were shown to be sensitive to inhibition by PatA (Low et al., 2005), we selected RKO cells (IC50 of 0.4 nM in cell proliferation assay) to prepare lysates for the isolation and identification of target protein(s). We often select RKO cells for target identification of small molecules using biotin-conjugates, because they appear to be particularly suitable for target protein isolation. 

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