Synthesis and Screening of a Combinatorial Peptide Library for Ligands to Target Transferrin: Miniaturizing the Library
Description: Combinatorial libraries are used in the search for ligands that bind to target proteins. Fmoc solid-phase peptide synthesis is routinely used to generate such libraries. Microwave-assisted peptide synthesis was employed here to decrease reaction times by 80-90%. Two One-Bead-One-Compound combinatorial libraries were synthesized on 130μm beads (one containing 750 members and the other 16, 807). The use of smaller solid supports would have many important practical advantages including; increased library diversity per unit mass, smaller quantities of library needed to generate hits, and screening could be conducted by using a standard flow cytometer. To this end, a miniaturized peptide library was synthesized on 20 μm beads to demonstrate proof of principle. A small sample from the 16,807-member library was screened against transferrin-AlexaFluro 647, a protein responsible for iron transport in vivo. A number of hits were identified and sequenced using techniques coupling nanomanipulation with nanoelectrospray mass spectrometry.
Date: August 2010
Creator: Brown, Jennifer Marie
Item Type: Thesis or Dissertation