Characterization of self- and hetero-interactions of bacterial cell division proteins FtsZ and sZipA by composition gradient – static light scattering (CG-SLS)

8 páginas, 7 figuras -- PAGS nros. 10780-10787We have characterized the self-association of FtsZ in its GDP-bound state (GDP-FtsZ) and the heteroassociation of FtsZ and a soluble recombinant ZipA (sZipA) lacking the N-terminal transmembrane domain by means of composition gradient−static light scattering (CG−SLS) and by measurement of sedimentation equilibrium. CG−SLS experiments at high ionic strengths and in the presence of 5 mM Mg2+ show that, while FtsZ self-associates in a noncooperative fashion, sZipA acts as a monomer. CG−SLS data obtained from mixtures of FtsZ (A) and sZipA (B) in the presence of Mg2+ are quantitatively described by an equilibrium model that takes into account significant scattering contributions from B, A1, A2, A3, A4, A5, A6, A1B, A2B, A3B, and A4B. However, in the absence of Mg2+ (with EDTA), the data are best explained by an equilibrium model in which only B, A1, A2, A3, A1B, and A2B contribute significantly to scattering. The best-fit molecular weights of monomeric A and B are in good agreement with values calculated from amino acid composition and with values obtained from sedimentation equilibrium. The latter technique also confirmed the interaction between sZipA and GDP-FtsZ. Moreover, the association model that best describes the CG−SLS data is in qualitative agreement with the sedimentation data. From these results, it follows that the binding of sZipA to GDP-FtsZ is of moderate affinity and does not significantly affect the interactions between FtsZ monomers. Under the working conditions used, only one sZipA binds to FtsZ oligomers with a length of six at most. The observed behavior would be compatible with FtsZ fibrils being anchored in vivo to the bacterial inner plasma membrane by substoichiometric binding of membrane-bound ZipAThis work was supported by Plan Nacional (Ministerio de Ciencia e Innovación, Spain) Grants BIO2008-04478-C03 to G.R. Grant DIVINOCELL FP/HEALTH-F3-2009-223432 (European Commision) to G.R., and Grant COMBACT S-BIO-0260/2006 (Comunidad de Madrid) to J.M. and G.R. A.M. is a predoctoral fellow from the Ministerio de Ciencia e Innovación of Spain. P.L.-N. was a recipient of a predoctoral fellowship from the Comunidad de Madrid. The research of A.P.M. is supported by the Intramural Research Program of the National Institute of Diabetes and Digestive and Kidney Diseases. J.M. was the recipient of a Ramón y Cajal fellowship financed by the Spanish Ministerio de Ciencia e Innovación and the European Social FundPeer reviewe