Structure and encapsulation of carbonic anhydrase within the α-carboxysome

TL;DR

This study reveals how carbonic anhydrase is structured and encapsulated within α-carboxysomes, offering insights for synthetic biology applications.

Contribution

The study identifies a trimer-of-dimers structure of α-carboxysomal carbonic anhydrase and its interaction with carboxysome shells.

Findings

HnCsoSCA forms a trimer-of-dimers structure without a zinc ion at the hexameric interface.

HnCsoSCA interacts with the CsoS1A shell hexamer and bridges the shell and Rubisco.

HnCsoSCA is incorporated into synthetic minishells at the inner surface without requiring CsoS2.

Abstract

Carbonic anhydrase plays a pivotal role in biological carbon capture by catalyzing the interconversion of CO2 and bicarbonate. This study provides insights into the structure and encapsulation of carbonic anhydrase (CA) within the α-carboxysome, a proteinaceous organelle for CO2 fixation. The results reveal that α-carboxysomal CA from the chemoautotrophic, nonphotosynthetic bacterium Halothiobacillus neapolitanus (HnCsoSCA) forms a trimer-of-dimers structure in the absence of a zinc ion at the hexameric interface. Using synthetic α-carboxysome shells, we show that HnCsoSCA interacts with the CsoS1A shell hexamer and bridges the shell facet and Rubisco. These findings advance our understanding of α-carboxysome assembly and encapsulation mechanisms, paving the way for potential applications of carboxysome structures in synthetic biology and biotechnology.

Carboxysomes in cyanobacteria and certain proteobacteria enable efficient CO2 fixation by encapsulating ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) and carbonic anhydrase (CA) within a semipermeable shell. Sequestered CA catalyze the rapid interconversion of CO2 and HCO3−, supplying elevated levels of CO2 to boost Rubisco carboxylation. Despite its essential role, the structure and encapsulation of CA within carboxysomes remain poorly understood. Here, we determined the molecular structure of α-carboxysomal CA from the model chemoautotrophic bacterium Halothiobacillus neapolitanus (HnCsoSCA). HnCsoSCA adopts a trimer-of-dimers oligomeric structure without the incorporation of a zinc ion at its symmetric center. Using synthetic minishells, we demonstrate that HnCsoSCA interacts with the CsoS1A shell hexamer and is incorporated into the minishells at the inner surface, independent of the CsoS2 linker protein. HnCsoSCA truncations suggest nonspecific interactions between HnCsoSCA and CsoS1A. We further show that HnCsoSCA bridges Rubisco and the shell facets. Our study offers insights into the assembly and encapsulation mechanisms of α-carboxysomes and provides the framework for reprogramming carboxysome structures for synthetic biology and biotechnological applications.

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