Identification of cytosolic sialidase NEU2 associated proteins bt mass spectrometric analysis

Abstract

Sialidases (Neuraminidases) are the enzymes which remove sialic acids from glycoproteins, oligosacharides and glycolipids. Four mammalian sialidases have been identified which are lysosomal sialidase (Neu1), cytosolic sialidase (Neu2), plasma membrane sialidase (Neu3), and mitochondrial/lysosomal sialidase (Neu4). These enzymes differ in their subcellular localizations, expression levels in different cells and tissues, substrate specificities and optimum pH levels. Cytosolic Neu2 enzyme has an active role on a wide range of subtances including oligosaccharides, glycopeptides and gangliosides. Studies on the Neu2 enzyme also showed that this enzyme is involved in different cellular events including cancer metabolism, neuronal differentiation and myoblast differentiation, proliferation and hypertrophy. However, it has not been shown whether Neu2 interacts with other proteins within the cell. Therefore, in this study we aimed to identify Neu2 associated proteins by using InterPlay Mammalian TAP System and ESI-LC-MS/MS analysis. Proteins in the Neu2 protein complex were identified by three different database search software such as PGLS, Mascot and X!Tandem. As a result of experiment Actin proteins (Alpha Actin, Gamma Actin and Beta Actin), and Calsyntenin-2 protein were found as a candidate protein for Neu2 association. The interaction between Neu2 and β-Actin proteins was confirmed by western blot analysis.