DATE: Thursday, 20 January 2005

SPEAKER: Dr. Haleem Issaq

TOPIC: Effective Fractionation and Separation Strategies for Proteomic Analysis

TIME: 7:30 pm

LOCATION: Shimadzu Scientific Inc. Training Center, 7100 Riverwood Drive, Columbia, MD 21406

Dinner will be at 5:30 pm at the Rams Head Tavern in Savage, MD. If you plan to attend the dinner, please send an email to reservations@wcdg.org

ABSTRACT:

Effective Fractionation and Separation Strategies for Proteomic Analysis Haleem Issaq, Thomas P. Conrads and Timothy D. Veenstra

The proteome describes the entire compliment of proteins expressed by a cell at a point in time. The analysis of a proteome, typically involves the digestion of the intact proteins followed by fractionation using a variety of chromatographic and/or electrophoretic techniques prior to their actual identification using mass spectrometry. Significant research has been carried out to develop electrophoretic and chromatographic methods for the resolution and identification of all peptides (proteins) in a given proteome. A serum proteome may contain up to 20,000 proteins with a concentration dynamic range of 10 10 which when digested may result in 600,000 – 900,000 peptides. No single chromatographic or electrophoretic procedure to date possesses the peak capacity required to resolve such a complex mixture into its individual components. Many attempts have been made using different electrophoretic and chromatographic modes and a combination of both techniques, off-line and on-line, to separate such complex protein/peptide mixtures. However, no method presently exists that can be used to separate, detect and quantify all proteins within a given proteome. Fractionation of a proteome digest is a critical aspect of mass spectral identification of peptides. Conventional MS/MS cycle times for most instruments typically enable only a limited number of these peptides to be selected for collision induced dissociation (CID). Therefore, limiting the number of co-eluting peptides helps to increase the percentage of species that may be identified. The aim of this presentation is to highlight the different liquid phase separation strategies that have been employed for fractionation and separation of complex mixtures of peptides. Also, mass spectrometric approaches, for maximizing the number of peptide identifications, such as gas phase fractionation and repeat injections will be discussed. Acknowledgement: Supported by NCI Contract No. NO1-CO-12400.

BIOGRAPHICAL SKETCH

Dr. Haleem J. Issaq is a Principal Scientist and Director of the Separation Technologies Laboratory at SAIC Frederick, NCI-Frederick. He received his B. Sc. From the American college of Istanbul in 1965, M.Sc. from the Technion in 1968 and a Ph.D. in analytical chemistry from Georgetown University in 1972 and joined the Frederick Cancer Research and Development Center the same year. He has been in charge of separation sciences dealing with the separation, detection and quantitation of analytes from complex biological and environmental mixtures using thin layer chromatography, gas chromatography, high performance liquid chromatography and capillary electrophoresis. The complexity of the analytes have ranged from metals, to carcinogens, to natural products, to proteins and DNA fragments. His current interests are in developing multidimensional separation schemes for complex mixtures such as human serum proteome. He has published extensively (over 245 publications) on atomic absorption spectroscopy, thin layer chromatography, high pressure liquid and gas chromatography, and capillary electrophoresis. He is the editor of “A Century of Separation Science” which was published in 2002. He is the recipient of the 1987 Eastern Analytical Symposium Chromatography Award, and Maryland Chemist of the Year 2000 award by ACS. Dr. Issaq is Founder and General Chairman of the Annual Frederick Conference on Capillary Electrophoresis, which is in its 15th year, Chairman and Organizer of the first three American-Hungarian Chromatography Symposium, Program Chairman and Member of the Program Committee and Governing Board of the Eastern Analytical Symposium from 1984-1999, Associate Editor, Journal of Liquid Chromatography, Guest Editor, Journal of Chromatography, J. of Separation Science, and Electrophoreis. He is a member of the editorial boards of the Journal of Capillary Electrophoresis, Electrophoresis, and Instrumentation Science and Technology. Dr. Issaq is the recipient of the Maryland Chemist of the year award, the “1998 Neuron Award” presented by the Director of the Institute for Basic Research in Developmental Disabilities and the Governor of the State of New York, and the 1998 Distinguished Career Service Award presented by SAIC Frederick, A Division of Science Applications International Corporation, and his group was awarded the 1999 Outstanding Science Achievement Award by SAIC Frederick.