Overexpression and Characterization of The C-Terminal Domain of Human Siva1, A Proapoptotic Factor and Cytoskeleton Binding Protein.
Siva1 protein interacts with tumor protein p53 and with the member of the tumor necrosis factor receptor superfamily, stathmin, among others. These proteins are related to several pathways involved in cancer and are therefore strong candidate targets for drug design. This study aimed to characterize the biophysical properties of Siva 1 C- terminal domain to contribute to the discovery of new target directed drugs. Siva1 protein interacts with tumor protein p53 and with the member of the tumor necrosis factor receptor superfamily, stathmin, among others. These proteins are related to several pathways involved in cancer and are therefore strong candidate targets for drug design. This study aimed to characterize the biophysical properties of Siva 1 C- terminal domain to contribute to the discovery of new target directed drugs. The C-terminus Siva1 domain (residues 84-175) was fused to glutathione Stransferase (GST) and expressed in an E coli system and the recombinant GST-Siva C-terminus was purified by GSTTagged Protein affinity and gel filtration chromatography. We tested the biological activity of the purified Siva Cterminus domain in a Jurkat extract cell line and found that the protein interacted with natural binders. Biophysical and biochemical assays have demonstrated monodispersion of the protein in solution with a predominant unfolded and elongated shape. However, at high concentrations, the protein showed a tendency to form soluble aggregates. These results are expected to lead to further progress in the understanding of Siva1 properties and target-directed drug design.
Larissa E C
LE
Dantas
Hemocentro, Faculty of Medical Sciences at University of Campinas, Rua Carlos Chagas, 480; Cidade Universitária Zeferino Vaz; 13083-878; Campinas, São Paulo, Brasil. sara@unicamp.br.
Sara T O
ST
Saad
Carlos H I
CH
Ramos
Serge
S
Bénichou
D051017:Apoptosis Regulatory Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D006801:Humans; D019169:Jurkat Cells; D011485:Protein Binding; D017510:Protein Folding; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D051017:Apoptosis Regulatory Proteins; D011993:Recombinant Fusion Proteins; C106549:SIVA1 protein, human; D005982:Glutathione Transferase