Crystallographic Insights into the Pore Structures and Mechanisms of the EutL and EutM Shell Proteins of the Ethanolamine-Utilizing Microcompartment ofEscherichia coli

Abstract

The ethanolamine-utilizing bacterial microcompartment (Eut-BMC) of Escherichia coli is a polyhedral organelle that harbors specific enzymes for the catabolic degradation of ethanolamine. The compartment is composed of a proteinaceous shell structure that maintains a highly specialized environment for the biochemical reactions inside. Recent structural investigations have revealed hexagonal assemblies of shell proteins that form a tightly packed two-dimensional lattice that is likely to function as a selectively permeable protein membrane, wherein small channels are thought to permit controlled exchange of specific solutes. Here, we show with two nonisomorphous crystal structures that EutM also forms a two-dimensional protein membrane. As its architecture is highly similar to the membrane structure of EutL, it is likely that the structure represents a physiologically relevant form. Thus far, of all Eut proteins, only EutM and EutL have been shown to form such proteinaceous membranes. Despite their similar architectures, however, both proteins exhibit dramatically different pore structures. In contrast to EutL, the pore of EutM appears to be positively charged, indicating specificity for different solutes. Furthermore, we also show that the central pore structure of the EutL shell protein can be triggered to open specifically upon exposure to zinc ions, suggesting a specific gating mechanism.

Bioinformatic Characterization of Glycyl Radical Enzyme-Associated Bacterial Microcompartments

“…BMC-T proteins, which form trimeric pseudohexamers, are considered relatively minor components of BMC shell facets. Structures of some BMC-Ts indicate that the residues surrounding the pores formed by the trimers can have alternate conformations to gate the pores (18,(68)(69)(70)(71). A subset of BMC-Ts forms a distinct clade, and these pseudohexamers stack and are gated (68,69).…”

Eut Bacterial Microcompartments: Insights into Their Function, Structure, and Bioengineering Applications

“…2 ). It is interesting to note that under different crystallization conditions, packing of EutM monomers within the crystal lattice is such that a 2D protein array of hexamers can be formed [Takenoya et al, 2010], suggesting a major role in self-assembly of the facets of the microcompartment shell. The properties of either side of the hexameric assembly are markedly different; one side can be described as concave, displaying an obvious depression, which is typically the side where the N-and C-termini are located.…”

“…EutM and CD1918 [Pitts et al, 2012;Takenoya et al, 2010;Tanaka et al, 2008] display a more open pore than that of EutS, with a diameter of 9 Å, and a positively charged pore surrounded by positively charged patches, which has been suggested to play a role in transport of the acetyl-phosphate product of Eut ( fig. 3 ).…”

“…One example of a tandem BMC domain is that of EutL, which is formed by two circularly permuted domains [Takenoya et al, 2010;Tanaka et al, 2010]. EutL has been crystallized in two distinct forms, with significant conformational changes that lead to an obvious opening and closing of the central pore, which in its open form has a radius of 11 Å.…”

“…The BMC domain, which is conserved across all types of BMC, typically consists of 90 amino acids, adopts an α/β barrel fold, and has a central four-stranded antiparallel sheet that is flanked by alpha helices, as exemplified by the crystal structure of EutM from E. coli and the EutM homologue CD1918 from C. difficile [Pitts et al, 2012;Takenoya et al, 2010;. Monomers of the BMC domain self-assemble to form cyclic hexamers; each hexamer represents a single tile in the regular mosaic pattern of each facet of the microcompartment shell ( fig.…”