Drosophila phosrestin I in vivo determined by mass spectrometric analysis of proteins separated by two-dimensional gel electrophoresis



Full-text Article (Subscribers only)
Full-text article
(subscribers only)

Search
Search

Go Back

Eur. J. Mass Spectrom. 3, 367 - 378 (1997)

The phosphorylation site and desmethionyl N-terminus of Drosophila phosrestin I in vivo determined by mass spectrometric analysis of proteins separated by two-dimensional gel electrophoresis

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.

ABSTRACT:
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.

Keywords: electrospray ionization (ESI), LC-MS, collision-induced dissociation (CID), two-dimensional electrophoresis (2-Dgel), in-gel digestion, protein modification, phosrestin I, phosphorylation, Drosophila.


© IM Publications
Any problems? E-mail .