AVS 53rd International Symposium
    Biomaterial Interfaces Tuesday Sessions
       Session BI-TuP

Paper BI-TuP19
Metal Ion Impregnated PMMA as On-Probe Affinity Capture MALDI (OPAC-MALDI) Probes for the Fractionation of Protein Mixtures

Tuesday, November 14, 2006, 6:00 pm, Room 3rd Floor Lobby

Session: Biomaterial Interfaces Poster Session
Presenter: G.S. Fernando, Southern Illinois University
Authors: G.S. Fernando, Southern Illinois University
G.R. Kinsel, Southern Illinois University
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MALDI Mass Spectrometry (MS) is currently broadly used in the structural characterization and identification of proteins and other biological molecules. Fractionation of proteins into groups according to their physical or chemical properties is usually an important first step before MALDI MS analysis. Conventional methods typically involve external sample manipulation which limits the overall throughput of the analysis. Incorporation of these fractionation steps directly on a MALDI target plate has given rise to various novel protein chip technologies. In the present studies, the use of a metal impregnated, commercially available, comparatively inexpensive and easily fabricated, poly (methyl methacrylate) (PMMA) surface for the separation of metal binding proteins is explored. Identification of metal binding proteins is expected to facilitate the evaluation of the function of metal ions in protein folding, assembly, stability, conformational change, and catalysis. To produce the chemically modified MALDI target, PMMA is first hydrolyzed with 3M NaOH overnight to obtain carboxylic acid functional groups on the surface. After thorough washing with MilliQ water, the carboxyl groups are further reacted with metal ions, such as Cu2+, Zn2+ or Ni2+, by incubating the PMMA probes in a metal salt solution overnight. All surface modification steps are characterized by XPS, FTIR, AFM, MALDI MS and contact angle measurements and compared with unmodified PMMA. These metal incorporated PMMA MALDI targets are shown to selectively bind phosphorylated proteins, histidine rich proteins and proteins with terminal arginine residues in a series of control studies. Subsequently, the PMMA targets are further evaluated for their ability to fractionate complex mixtures of proteins derived from bacterial sources.