Cloning,Overexpression, Purification and Preliminary Characterization of Human Septin 8 |
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Authors: | T A C B Souza J A R G Barbosa |
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Institution: | 1.Institute of Biology,State University of Campinas (UNICAMP),Campinas,Brazil;2.Brazilian Synchrotron Light Laboratory (LNLS),Campinas,Brazil;3.Pós-Gradua??o em Ciências Gen?micas e Biotecnologia,Universidade Católica de Brasília,Brasília,Brazil |
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Abstract: | Mammalian septins comprise a family of 14 genes that encode GTP-binding proteins involved in important cellular processes
such as cytokinesis and exocytosis. Expression of three different constructs encoding human septin 8 were analyzed and the
results show that SEPT8GC, a clone expressing the conserved domain plus C-terminal domain of human septin 8 yields the highest
amount of recombinant protein. This protein was purified by affinity chromatography followed by a gel filtration chromatography.
CD spectrum of SEPT8GC is characteristic of folded proteins and it presents a transition profile with a T
m of 54 °C. Fluorescence emission spectra, analytic gel filtration and DLS reflect the sample oligomeric heterogeneity with
the predominance of dimers in solution. Homology models indicate clearly that the preferred dimer interface is the one comprising
the GTP binding site. |
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Keywords: | |
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