Bacterial microcompartment
Bacterial microcompartments are organelle-like structures found in bacteria. They consist of a protein shell that encloses enzymes and other proteins. BMCs are typically about 40–200 nanometers in diameter and are made entirely of proteins. The shell functions like a membrane, as it is selectively permeable. Other protein-based compartments found in bacteria and archaea include encapsulin nanocompartments and big gas vesicles.
Discovery
The first BMCs were observed in the 1950s in electron micrographs of cyanobacteria, and were later named carboxysomes after their role in carbon fixation was established. Until the 1990s, carboxysomes were thought to be an oddity confined to certain autotrophic bacteria. But then genes coding for proteins homologous to those of the carboxysome shell were identified in the pdu and eut operons. Subsequently, transmission electron micrographs of Salmonella cells grown on propanediol or ethanolamine showed the presence of polyhedral bodies similar to carboxysomes. The term metabolosome is used to refer to such catabolic BMCs.Although the carboxysome, propanediol utilizing, and ethanolamine utilizing BMCs encapsulate different enzymes and therefore have different functions, the genes encoding for the shell proteins are very similar. Most of the genes from experimentally characterized BMCs are located near one another in distinct genetic loci or operons. There are currently over 20,000 bacterial genomes sequenced, and bioinformatics methods can be used to find all BMC shell genes and to look at what other genes are in the vicinity, producing a list of potential BMCs. In 2014, a comprehensive survey identified 23 different loci encoding up to 10 functionally distinct BMCs across 23 bacterial phyla. In 2021, in an analysis of over 40,000 shell protein sequences, it was shown that at least 45 phyla have members that encode BMCs, and the number of functional types and subtypes has increased to 68. The role of BMCs in the human microbiome is also becoming clear.
Shells
Protein families forming the shell
The BMC protein shell appears icosahedral or quasi-icosahedral, and is formed by hexameric and pentameric protein subunits. Structures of intact shells have been determined for three functionally distinct BMC types: carboxysomes, the GRM2 organelles involved in choline catabolism, and a metabolosome of unknown function. Collectively, these structures shown that the basic principles of shell assembly are universally conserved across functionally distinct BMCs.The BMC shell protein family
The major constituents of the BMC shell are proteins containing Pfam00936 domain. These proteins form oligomers that are hexagonal in shape and form the facets of the shell.Single-domain proteins (BMC-H)
The BMC-H proteins, which contain a single copy of the Pfam00936 domain, are the most abundant component of the facets of the shell. The crystal structures of a number of these proteins have been determined, showing that they assemble into cyclical hexamers, typically with a small pore in the center. This opening is proposed to be involved in the selective transport of the small metabolites across the shell. Most BMCs contain multiple distinct types of BMC-H proteins that tile together to form the , likely reflecting the range of metabolites that must enter and exit the shell.Tandem-domain proteins (BMC-T)
A subset of shell proteins are composed of tandem copies of the Pfam00936 domain, this evolutionary event has been recreated in the lab by the construction of a synthetic BMC-T protein. Structurally characterized BMC-T proteins form trimers that are pseudohexameric in shape. Some BMC-T crystal structures show that the trimers can stack in a face-to-face fashion. In such structures, one pore from one trimer is in an "open" conformation, while the other is closed – suggesting that there may be an airlock-like mechanism that modulates the permeability of some BMC shells. This gating appears to be coordinated across the surface of the shell. Another subset of BMC-T proteins contain a cluster, and may be involved in electron transport across the BMC shell. Metal centers have also been engineered into BMC-T proteins for conducting electrons.The EutN/CcmL family (BMC-P)
Twelve pentagonal units are necessary to cap the vertices of an icosahedral shell. Crystal structures of proteins from the EutN/CcmL family have been solved and they typically form pentamers. The importance of the BMC-P proteins in shell formation seems to vary among the different BMCs. It was shown that they are necessary for the formation of the shell of the PDU BMC as mutants in which the gene for the BMC-P protein was deleted cannot form shells, but not for the alpha-carboxysome: without BMC-P proteins, carboxysomes will still assemble and many are elongated; these mutant carboxysomes appear to be "leaky".Evolution of BMCs and relation to viral capsids
While the BMC shell is architecturally similar to many viral capsids, the shell proteins have not been found to have any structural or sequence homology to capsid proteins. Instead, structural and sequence comparisons suggest that both BMC-H and BMC-P, most likely, have evolved from bona fide cellular proteins, namely, PII signaling protein and OB-fold domain-containing protein, respectively.Permeability of the shell
It is well established that enzymes are packaged within the BMC shell and that some degree of metabolite and cofactor sequestration must occur. However, other metabolites and cofactors must also be allowed to cross the shell in order for BMCs to function. For example, in carboxysomes, ribulose-1,5-bisphosphate, bicarbonate, and phosphoglycerate must cross the shell, while carbon dioxide and oxygen diffusion is apparently limited. Similarly, for the PDU BMC, the shell must be permeable to propanediol, propanol, propionyl-phosphate, and potentially also vitamin B12, but it is clear that propionaldehyde is somehow sequestered to prevent cell damage. There is some evidence that ATP must also cross some BMC shells.It has been proposed that the central pore formed in the hexagonal protein tiles of the shell are the conduits through which metabolites diffuse into the shell. For example, the pores in the carboxysome shell have an overall positive charge, which has been proposed to attract negatively charged substrates such as bicarbonate. In the PDU microcompartment, mutagenesis experiments have shown that the pore of the PduA shell protein is the route for entry of the propanediol substrate. For larger metabolites, a gating mechanism in some BMC-T proteins is apparent. In the EUT microcompartment, gating of the large pore in the EutL shell protein is regulated by the presence of the main metabolic substrate, ethanolamine.
The presence of iron-sulfur clusters in some shell proteins, presumably in the central pore, has led to the suggestion that they can serve as a conduit through which electrons can be shuttled across the shell.
Types
Comprehensive surveys of microbial genome sequence data indicated more than 60 different metabolic functions encapsulated by BMC shells. The majority are involved in either carbon fixation or aldehyde oxidation. A webserver, BMC Caller, allows identification of the BMC type based on the protein sequences of the BMC locus components.Carboxysomes: carbon fixation
Carboxysomes encapsulate ribulose-1,5-bisphosphate carboxylase/oxygenase and carbonic anhydrase in -fixing bacteria as part of a concentrating mechanism. Bicarbonate is pumped into the cytosol and diffuses into the carboxysome, where carbonic anhydrase converts it to carbon dioxide, the substrate of RuBisCO. The carboxysome shell is thought to be only sparingly permeable to carbon dioxide, which results in an effective increase in carbon dioxide concentration around RuBisCO, thus enhancing fixation. Mutants that lack genes coding for the carboxysome shell display a high requiring phenotype due to the loss of the concentration of carbon dioxide, resulting in increased oxygen fixation by RuBisCO. The shells have also been proposed to restrict the diffusion of oxygen, thus preventing the oxygenase reaction, reducing wasteful photorespiration.Metabolosomes: aldehyde oxidation
In addition to the anabolic carboxysomes, several catabolic BMCs have been characterized that participate in the heterotrophic metabolism via short-chain aldehydes; they are collectively termed metabolosomes.In 2014 it was proposed that the despite their functional diversity, the majority of metabolosomes share a common encapsulated chemistry driven by three core enzymes: aldehyde dehydrogenase, alcohol dehydrogenase, and phosphotransacylase. Because aldehydes can be toxic to cells and/or volatile, they are thought to be sequestered within the metabolosome. The aldehyde is initially fixed to coenzyme A by a NAD+-dependent aldehyde dehydrogenase, but these two cofactors must be recycled, as they apparently cannot cross the shell. These recycling reactions are catalyzed by an alcohol dehydrogenase, and a phosphotransacetylase, resulting in a phosphorylated acyl compound that can readily be a source of substrate-level phosphorylation or enter central metabolism, depending on if the organism is growing aerobically or anaerobically. It seems that most, if not all, metabolosomes utilize these core enzymes. Metabolosomes also encapsulate another enzyme that is specific to the initial substrate of the BMC, that generates the aldehyde; this is the defined signature enzyme of the BMC.