A robust synthetic biology toolkit to advance carboxysome study and redesign.

Carboxysomes are polyhedral protein organelles that microorganisms use to facilitate carbon dioxide assimilation. They are composed of a modular protein shell which envelops an enzymatic core mainly comprised of physically coupled Rubisco and carbonic anhydrase. While the modular construction principles of carboxysomes make them attractive targets as customizable metabolic platforms, their size and complexity can be a hinderance.

An invariant C-terminal tryptophan in McdB mediates its interaction and positioning function with carboxysomes.

Bacterial microcompartments (BMCs) are widespread, protein-based organelles that regulate metabolism. The model for studying BMCs is the carboxysome, which facilitates carbon fixation in several autotrophic bacteria. Carboxysomes can be distinguished as type α or β, which are structurally and phyletically distinct.

Positioning of cellular components by the ParA/MinD family of ATPases.

The ParA/MinD (A/D) family of ATPases spatially organize an array of genetic- and protein-based cellular cargos across the bacterial and archaeal domains of life. By far, the two best-studied members, and family namesake, are ParA and MinD, involved in bacterial DNA segregation and divisome positioning, respectively. ParA and MinD make protein waves on the nucleoid or membrane to segregate chromosomes and position the divisome.

An experimental framework to assess biomolecular condensates in bacteria.

High-resolution imaging of biomolecular condensates in living cells is essential for correlating their properties to those observed through in vitro assays. However, such experiments are limited in bacteria due to resolution limitations. Here we present an experimental framework that probes the formation, reversibility, and dynamics of condensate-forming proteins in Escherichia coli as a means to determine the nature of biomolecular condensates in bacteria.

Dissecting the phase separation and oligomerization activities of the carboxysome positioning protein McdB.

Across bacteria, protein-based organelles called bacterial microcompartments (BMCs) encapsulate key enzymes to regulate their activities. The model BMC is the carboxysome that encapsulates enzymes for CO fixation to increase efficiency and is found in many autotrophic bacteria, such as cyanobacteria. Despite their importance in the global carbon cycle, little is known about how carboxysomes are spatially regulated.