Bacterial microcompartments as metabolic modules for plant synthetic biology

Abstract

SUMMARYBacterial microcompartments (BMCs) are megadalton-sized protein assemblies that enclose segments of metabolic pathways within cells. They increase the catalytic efficiency of the encapsulated enzymes while sequestering volatile or toxic intermediates from the bulk cytosol. The first BMCs discovered were the carboxysomes of cyanobacteria. Carboxysomes compartmentalize the enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) with carbonic anhydrase. They enhance the carboxylase activity of RuBisCO by increasing the local concentration of CO 2 in the vicinity of the enzyme's active site. As a metabolic module for carbon fixation, carboxysomes could be transferred to eukaryotic organisms (e.g. plants) to increase photosynthetic efficiency. Within the scope of synthetic biology, carboxysomes and other BMCs hold even greater potential when considered a source of building blocks for the development of nanoreactors or three-dimensional scaffolds to increase the efficiency of either native or heterologously expressed enzymes. The carboxysome serves as an ideal model system for testing approaches to engineering BMCs because their expression in cyanobacteria provides a sensitive screen for form (appearance of polyhedral bodies) and function (ability to grow on air). We recount recent progress in the re-engineering of the carboxysome shell and core to offer a conceptual framework for the development of BMC-based architectures for applications in plant synthetic biology.

Bacterial microcompartments

“…Furthermore, a native RuBisCO-deletion tobacco plant line was able to grow when it was engineered so that a cyanobacterial RuBisCO could be produced in its chloroplasts; the addition of the gene that encodes for the SSLDs of CcmM led to the formation of a structure resembling a pro-carboxysome 124 . This first inter-kingdom heterologous expression of BMC components represents a leap towards the objective of utilizing carboxysomes in plants for enhanced photosynthesis in crops 25,62, 119,123–126 and, more generally, for building bacterial organelles within eukaryotic cells.…”

“…The same strategy that is based on encapsulation peptides could also be used to target synthetic scaffolding proteins 139 to the lumen of BMCs (Figure 5). This scaffold and the enzymes of the synthetic pathway would be engineered to specifically interact with each other, enabling the precise control of protein stoichiometry 25 .…”

“…Accordingly, a EUT AldDH was able to functionally replace the native enzyme of a PDU BMC 138 . This strategy could be extended to any core enzyme, by replacing one of the native enzymes with a catalytically distinct member with the same structural fold 7, 25, 118 . Surface features (for example, size and charge of specific amino acids) of the homologous enzyme could be altered to resemble those known to facilitate protein-protein interactions in the native core.…”

Harnessing transcription for bioproduction in cyanobacteria

“…As photosynthetic organisms, they are ideal for CO 2 assimilation and conversion to biochemicals, concurrently alleviating the rising levels of atmospheric CO 2 . Cyanobacteria contain specialized carbon‐concentrating compartments that increase carbon fixation efficiency, but can also be used as a scaffold to target biosynthetic enzymes to create specialized metabolic microcompartments (Gonzalez‐Esquer et al ). Moreover, cyanobacteria can photosynthetically generate and efficiently distribute redox power to fuel redox‐dependent heterologous pathways, due to the plasticity of the photosynthetic electron carrier proteins (Mellor et al ).…”

Synthetic biology for basic and applied plant research

“…They can encapsulate enzymes, concentrate substrates, and help in the avoidance of toxic products as Gonzalez-Esquer et al (2016) describe. Cyanobacteria address one key problem that all photosynthetic organisms encounter, the natural inefficiency of the carbon-fixing enzyme RubBisCO, by encapsulating this enzyme in carboxysomes, which increases the local concentration of CO 2 around the enzyme.…”