Cloning, Overexpression, Purification and Preliminary Characterization of Human Septin 8

Author(s)

Souza, T. A. C. B. ^{[1, 2]} ; Barbosa, J. A. R. G. ^{[1, 3, 2]}

Affiliation

^[1] Univ Estadual Campinas, UNICAMP, Inst Biol, Campinas, SP - Brazil

^[2] Brazilian Synchrotron Light Lab LNLS, BR-13083970 Campinas, SP - Brazil

^[3] Univ Catolica Brasilia, BR-70790160 Brasilia, DF - Brazil

• Document type: Journal article
• Source: The Protein Journal; v. 29, n. 5, p. 328-335, JUL 2010.
• Web of Science Citations: 1

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 A degrees 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. (AU)

• FAPESP's process: 98/14138-2 - Center for Structural Molecular Biotechnology
• Grantee: Glaucius Oliva
• Support Opportunities: Research Grants - Research, Innovation and Dissemination Centers - RIDC