Lipoprotein Quantitation and Function Core

ABSTRACT (Core C) High quality, consistent and reproducible quantitation and functional data are critical for the success of this Program Project. The overarching goal of the HDL Quantitation and Function Core (Core C) is to meet this need in a centralized and consistent manner across the entire Program. The Core is structured to provide the following services: 1) quantitation of HDL particle size and concentration (whether isolated from plasma or reconstituted HDL particles) for specific structural studies, using calibrated differential ion mobility analysis (cIMA); 2) mass spectrometric analyses to characterize the protein composition of HDL particles isolated or prepared by Program investigators, and chemical crosslinking strategies to support their structural studies; the Core will also use mass spectrometry to confirm the identity and purity of the recombinant proteins prepared by Core D; 3) a combination of accurate measurement of HDL particle concentration by calibrated IMA and accurate quantification of apolipoproteins by LC-MS/MS, including quantification of the stoichiometry of apolipoproteins on HDL particles; 4) bioinformatic support for interpreting HDL-P and proteomic data sets and for combining the data with information from Gene Ontology, protein-protein interactions, and pathway analysis; 5) measurements of cholesterol and phospholipid efflux, using baby hamster kidney cells conditionally expressing WT and mutant human ABCA1. Core C will be led by Tomas Vaisar, Ph.D., working closely with Chongren Tang, Ph.D., and it will be located at the University of Washington South Lake Union campus, where Project 1 is also located. Both labs have extensive, well-documented expertise in all areas of the Core’s proposed services. In the previous PPG cycle, this Core performed over 1000 analyses of HDL-P and size, over 500 LC-MS/MS analyses of HDL’s proteome, and crosslinking studies for characterizing and experimentally verifying theoretical models of HDL and ABCA1 structures. These services will be extended in this proposal. Collectively, they will enable cost-effective and rigorous testing of the hypotheses generated by the various Projects.