UCSF

Bacterial Actin-Like Proteins (ALPs) participate in many biologically, clinically and commercially important processes, including segregation of low-copy plasmids. To better understand the biochemical principles underlying plasmid DNA segregation we purified a recently discovered plasmid-segregating ALP, Alp7A, and studied its structure and self-assembly in vitro. Monomeric Alp7A has a uniquely low affinity for nucleotides, binding ATP 1,000,000-fold more weakly than eukaryotic actin. The atomic structure of Alp7A (solved to 2.4 A resolution by x-ray crystallography) revealed that the low nucleotide affinity correlates with a unique nucleotide conformation. Alp7A has a low rate of spontaneous nucleation and, at low concentrations (<11 µM), forms ephemeral and highly dynamic polymers. At higher concentrations these ephemeral polymers are stabilized by lateral association, forming large bundles which contain antiparallel filaments. The accessory factor, Alp7R, dramatically increases the nucleation rate of Alp7A filaments and decreases their propensity to bundle, making them behave much more like ParM filaments. These nucleation and stabilization activities of Alp7R occur both in vitro and in vivo and do not require DNA, neither alp7C or non-specific DNA.