Structural and molecular genetic insight into a widespread sulfur oxidation pathway

Many sulfur-oxidizing bacteria store sulfur as water-insoluble globules during metabolic processes. While the Dsr protein family is known to be involved in sulfur oxidation, the exact roles of individual Dsr proteins remain unclear. Prior research has shown that sulfur globules are common in photo- and chemolithoautotrophic bacteria. However, the mechanisms by which these proteins function together are not fully understood. Some studies suggest that Dsr proteins may form a sulfur relay system. Yet, the precise sequence of reactions and the functional importance of specific residues are still unknown. This gap motivated further investigation into the Dsr system in Allochromatium vinosum. The lack of detailed structural and genetic data has limited progress in this area. Understanding these interactions could clarify sulfur oxidation pathways in diverse bacterial species.

This study aimed to clarify the functional roles of DsrEFH in sulfur oxidation within Allochromatium vinosum. The researchers focused on the DsrE and DsrH subunits of DsrEFH and their interaction with DsrC. They sought to determine whether DsrEFH is essential for sulfur degradation in sulfur globules. The study also aimed to identify conserved residues critical for protein interactions. Structural analysis of DsrEFH was another key goal. The team wanted to determine if DsrEFH interacts with DsrC in a manner similar to TusE and TusBCD systems. The investigation also aimed to compare the Dsr system with known sulfur relay systems in other bacteria. The ultimate purpose was to provide molecular evidence for sulfur transfer during oxidation.

The researchers used a deletion mutant of dsrE in Allochromatium vinosum to test the role of DsrEFH in sulfur oxidation. Complementation experiments were performed to restore sulfur degradation ability. The crystal structure of DsrEFH was determined using X-ray crystallography at 2.5 Å resolution. Phylogenetic analysis was conducted to identify conserved active sites in DsrE and DsrH. The team compared the structure of DsrEFH with known sulfur relay proteins like TusBCD and TusE. Protein–protein interactions were analyzed using biochemical assays. The conserved Cys78 in DsrE was mutated to assess its role in DsrC binding. The study also examined the role of Cys20 in DsrH in these interactions.

The DeltadsrE mutant was unable to degrade sulfur globules, confirming DsrEFH's essential role. Complementation with dsrEFH restored sulfur oxidation, proving its necessity. The crystal structure of DsrEFH revealed an alpha2beta2gamma2 configuration. Two distinct active site types were identified in DsrE and DsrH through phylogenetic analysis. Cys78 in DsrE is conserved and matches active cysteines in E. coli YchN and TusD. DsrEFH interacts with DsrC, a TusE homologue, via the conserved penultimate cysteine in DsrC’s C-terminus. Cys78 in DsrE is strictly required for DsrC binding, but Cys20 in DsrH is not. These findings suggest sulfur transfer occurs during oxidation via the Dsr system.

The study confirms that DsrEFH is essential for sulfur oxidation in Allochromatium vinosum. The structure and function of DsrEFH were clarified through complementation and crystallography. Cys78 in DsrE is critical for interaction with DsrC, indicating its role in sulfur transfer. DsrH’s Cys20 is not required for this interaction, suggesting functional redundancy. The conserved active sites in DsrE and DsrH are characteristic of sulfur-oxidizing bacteria. These findings support the hypothesis that sulfur transfer occurs during oxidation via the Dsr system. The study also highlights structural similarities between Dsr and Tus systems. The results provide a molecular framework for understanding sulfur oxidation in these bacteria.