Analysis of FtsZ Assembly by Light Scattering and Determination of the Role of Divalent Metal Cations

Abstract

ABSTRACT FtsZ is an ancestral homologue of tubulin that polymerizes in a GTP-dependent manner. In this study, we used 90° angle light scattering to investigate FtsZ polymerization. The critical concentration for polymerization obtained by this method is similar to that obtained by centrifugation, confirming that the light scattering is proportional to polymer mass. Furthermore, the dynamics of FtsZ polymerization could be readily monitored by light scattering. Polymerization was very rapid, reaching steady state within 30 s. The length of the steady-state phase was proportional to the GTP concentration and was followed by a rapid decrease in light scattering. This decrease indicated net depolymerization that always occurred as the GTP in the reaction was consumed. FtsZ polymerization was observed over the pH range 6.5 to 7.9. Importantly, Mg 2+ was not required for polymerization although it was required for the dynamic behavior of the polymers. It was reported that 7 to 25 mM Ca 2+ mediated dynamic assembly of FtsZ (X.-C. Yu and W. Margolin, EMBO J. 16:5455–5463, 1997). However, we found that Ca 2+ was not required for FtsZ assembly and that this concentration of Ca 2+ reduced the dynamic behavior of FtsZ assembly.