AVS 60th International Symposium and Exhibition
    Biomaterial Interfaces Thursday Sessions
       Session BI+AS+BA+NS+SS-ThA

Paper BI+AS+BA+NS+SS-ThA8
Determination of Orientation and Tertiary Structure of Adsorbed Protein on Material Surfaces by Chemical Modification and Peptide Mapping

Thursday, October 31, 2013, 4:20 pm, Room 102 B

Session: Biomolecules at Interfaces
Presenter: A.A. Thyparambil, Clemson University
Authors: A.A. Thyparambil, Clemson University
Y. Wei, Clemson University
R.A. Latour, Clemson University
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Chemical modification of targeted amino acid residues with peptide mapping via mass spectrometry (MS) is a promising technique to provide highly detailed information on the structural shifts and orientation of adsorbed protein by revealing adsorption-induced changes in amino acid solvent accessibility. A decrease in amino acid labeling (i.e., decreased solvent accessibility) is indicative of adsorbed orientation while an increase is indicative of tertiary unfolding. However, the potential of this method for the study of adsorbed protein structure is largely undeveloped at this time. The objective of our research was therefore to develop chemical modification and peptide mapping techniques that would help identify the dominant configuration of adsorbed protein on a material surface for a range of amino acid types. By directly comparing the extent of amino acid modification (profiles) from separate batch experiments targeting different types of amino acids, a fairly detailed picture of adsorption-induced changes in adsorbed protein structure can be obtained. In our current study, when unmodified segments of the protein without the targeted amino acid from the MS results was used as an internal standard for each of the batch experiments, a common baseline to directly compare the profiles of different amino acids could be obtained. Under these conditions, the configuration of hen egg white lysozyme (HEWL) when adsorbed on fused silica glass (glass), high density polyethylene (HDPE), and poly(methyl–methacrylate) (PMMA) was mapped by directly comparing the profiles of arginine (Arg), lysine (Lys), tryptophan (Trp), and carboxylic groups (Asp, Glu, C-terminus). For each of the targeted amino acid groups, the labeling procedure did not induce significant structural shifts, which was verified by circular dichroism spectropolarimetry. The resulting quantitative differences in the profiles of targeted amino acid residues in HEWL on different surfaces under different conditions correspond to different configuration of HEWL on each adsorbent surface. The developed technique has the potential for broad application and to be expanded to other targeted amino acids, thus providing highly detailed information on the adsorbed state of protein on any given surface.