Rebekah Gundry The Johns Hopkins University School of Medicine Office: 410-955-3022 FAX: 410-955-3420 E-mail: rebekahgundry@jhu.edu Job Title: Graduate Student Ph.D. in Analytical Chemistry (expected January, 2006) from The Johns Hopkins University

Speaker: Rebekah Gundry, Johns Hopkins University School of Medicine, Department of Pharmacology, Baltimore, MD 21205

Topic: Examination of the albumin-enriched fraction of human serum: protein separation, identification, and albumin-binding studies

Place: Building 549, Auditorium, NCI at Frederick, Frederick, MD

Time: Tuesday, November 8, 2005, at 2:00 PM

Abstract: Highly abundant serum proteins, such as albumin, are generally removed prior to proteomic analyses, as they mask the detection of lower abundance proteins. An inexpensive and rapid approach that employs chemical extraction for albumin depletion has recently been developed for proteomics applications, resulting in an albumin-depleted pellet and an albumin-enriched supernatant. The current work presents a method for characterizing the albumin-enriched fraction of human serum obtained via this chemical extraction procedure as part of the evaluation of this extraction method. The assessment of the albumin-enriched fraction included identifying the proteins/peptides in the albumin-enriched fraction, determining which, if any, of these are indeed albumin-binding proteins/peptides, and finally, assessing the reproducibility of the chemical extraction method in order to evaluate the potential for the albumin-enriched fraction to function as a subproteome for biomarker discovery. Understanding the proteins and or peptides that are bound to albumin is critical both with respect to biology, as this will change their free and/or active forms in the blood, as well as from the perspective of biomarker discovery. Bound proteins will or could have different clearance kinetics from their unbound forms. A total of 120 proteins were identified in this fraction, including both intact proteins and protein fragments. Three independent methods were compared to identify several albumin-binding proteins. Finally, combining the fact that this extraction method is reproducible, contains a number of reportedly potential biomarkers, and is less complex that human serum lends it potential to serve as a useful subproteomes for future biomarker studies. This rigorous assessment of the albumin-enriched fraction obtained from the chemical extraction method is a valuable asset when examining the proteins and peptides removed in the initial stages of a proteomics study and provides a model for the assessment of other albumin-depletion methods.