Peptide-functionalized polymeric nanoparticles for detecting Abl kinase activity
Abstract
The aim of this thesis project was to develop peptide-functionalized polymeric nanoparticles to be used as biosensors in monitoring Abl kinase activity in cells. The polyacrylamide nanoparticles were synthesized via a free radical water-in-oil microemulsion polymerization technique. Abltide (optimized artificial peptide substrate for Abl kinase) was labeled with bisacrylamide and polymerized with acrylamide monomers with (NP-2 and NP-3) and without (NP-1) cross-linkers. The rationale was to provide higher local concentration of the peptide at the particle surface to offer enhanced interaction with the kinase. NP-2 contained bisacrylamide cross-linkers while NP-3 contained SR415 (ethoxylated trimethylolpropane triacrylate esters) cross-linkers. Using dynamic light scattering (DLS) and transmission electron microscopy (TEM), the sizes of the nanoparticles were found to decrease with increasing concentration of AOT (surfactant). Using 15.5 as the molar ratio of water to surfactant, the sizes of all three variants of the nanoparticles were less than 100nm. The sensitivity of the nanoparticles to phosphorylation by Abl kinase was determined via in vitro kinase assays with HEK 293 cell lysates overexpressing GFP-Abl, and purified Abl kinase. Selected-reaction monitoring (SRM) and selected-ion monitoring (SIM) mass spectrometry were used as analytical detection techniques for the kinase assays
Degree
M.S.
Advisors
Parker, Purdue University.
Subject Area
Biochemistry|Polymer chemistry
Off-Campus Purdue Users:
To access this dissertation, please log in to our
proxy server.