Heterologous expression of human connexin 43 in Escherichia coli

Orateur : Soitenance de Thèse de Maria SILACHEVA (Institut de Biologie Structurale)

Membrane proteins are major functional components of biological membranes which are involved in key cellular processes. Connexins comprize a family of vertebrate integral MPs playing a central role in direct intercellular communication. They oligomerize and form cell-to-cell channels which cluster to gap junctions. Gap junctions mediate electrical and metabolic coupling between cells. We expressed connexin 43 and its truncated at residue 263 mutant in E. coli using (i) membrane targeting expression tag Mistic from B. subtilis (ii) connexin gene optimization for E. coli codon bias and minimization of free energy of mRNA secondary structure. Self-assembling of purified protein into hexamers was shown by SEC, DLS, and SAXS techniques. C-terminal cytoplasmic domain of connexin 43 (Cx43CT) and Mistic protein were overexpressed in E. coli and purified to homogeneity. Concentration-dependent oligomerization of Cx43CT was established.

Télécharger dans mon agenda