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Tomoya Kinumi, Sara L. Tobin and Hiroyuki Matsumoto*
Department of Biochemistry
and Molecular Biology, The University of Oklahoma Health Sciences Center, PO Box 26901, Oklahoma City, OK 73190, USA.
Kenneth W. Jackson
Department of
Medicine and Warren Medical Research Foundation, The University of Oklahoma Health Sciences Center, PO Box 26901, Oklahoma City, OK 73190, USA.
Mamoru
Ohashi
Department of Applied Physics and Chemistry, The University of Electro-Communications, Chofu, Tokyo 182, Japan. |
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Post-translational modifications of
proteins play crucial roles in modulating many cellular processes. In order to understand the physiological roles of post-translational protein modifications it is imperative to
determine the nature of the change in chemical structure involved in each protein modification. In our earlier work, we developed a method for the study of protein modification
through a streamlined combination of two-dimensional gel electrophoresis, in-gel digestion, high performance liquid chromatography and electrospray ionization tandem
quadrupole mass spectrometry. Using this method we determined the in vivo phosphorylation site of phosrestin I to be Ser366. Since our earlier work
described the method only briefly, we will present a full description of the method in this paper. In addition, by using this method, we also show that the N-terminus of phosrestin I is
desmethionylated in vivo. These techniques are easily adapted to the study of other proteins and will serve as powerful tools for the study of post-translational protein
modifications in general. |
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Keywords:
electrospray ionization (ESI), LC-MS, collision-induced dissociation (CID), two-dimensional electrophoresis (2-Dgel), in-gel digestion, protein
modification, phosrestin I, phosphorylation, Drosophila. |
