Akyıldız Demir, SeçilSeyrantepe, Volkan04.03. Department of Molecular Biology and Genetics04. Faculty of Science01. Izmir Institute of Technology2020-07-252020-07-2520190250-46851303-829Xhttps://doi.org/10.1515/tjb-2018-0089https://hdl.handle.net/11147/9567Background: Cytoplasmic sialidase (NEU2) plays an active role in removing sialic acids from oligosaccharides, gly-copeptides, and gangliosides in mammalian cells. NEU2 is involved in various cellular events, including cancer metabolism, neuronal and myoblast differentiation, proliferation, and hypertrophy. However, NEU2-interacting protein(s) within the cell have not been identified yet. Objective: The aim of this study is to investigate NEU2 interacting proteins using two-step affinity purification (TAP) strategy combined with mass spectrometry analysis. Methods: In this study, NEU2 gene was cloned into the pCTAP expression vector and transiently transfected to COS-7 cells by using PEI. The most efficient expression time of NEU2- tag protein was determined by real-time PCR and Western blot analysis. NEU2-interacting protein(s) were investigated by using TAP strategy combined with two different mass spectrometry experiment; LC-MS/MS and MALDI TOF/TOF. Results: Here, mass spectrometry analysis showed four proteins; a-actin, beta-actin, calmodulin and histone H1.2 proteins are associated with NEU2. The interactions between NEU2 and actin filaments were verified by Western blot analysis and immunofluorescence analysis. Conclusions: Our study suggests that association of NEU2 with actin filaments and other protein(s) could be important for understanding the biological role of NEU2 in mammalian cells.eninfo:eu-repo/semantics/closedAccessSialidaseNEU2ActinCalmodulinStreptavidinArticle2-s2.0-8507357438510.1515/tjb-2018-008910.1515/tjb-2018-0089