Author Correction: High-resolution cryo-EM structure of urease from the pathogen Yersinia enterocolitica.

A Correction to this paper has been published: https://doi.org/10.1038/s41467-020-19845-z .

High-resolution cryo-EM structure of urease from the pathogen Yersinia enterocolitica.

Urease converts urea into ammonia and carbon dioxide and makes urea available as a nitrogen source for all forms of life except animals. In human bacterial pathogens, ureases also aid in the invasion of acidic environments such as the stomach by raising the surrounding pH. Here, we report the structure of urease from the pathogen Yersinia enterocolitica at 2 Å resolution from cryo-electron microscopy. Y. enterocolitica urease is a dodecameric assembly of a trimer of three protein chains, ureA, ureB and ureC. The high data quality enables detailed visualization of the urease bimetal active site and of the impact of radiation damage. The obtained structure is of sufficient quality to support drug development efforts.

Cohesive Properties of the Caulobacter crescentus Holdfast Adhesin Are Regulated by a Novel c-di-GMP Effector Protein.

When encountering surfaces, many bacteria produce adhesins to facilitate their initial attachment and to irreversibly glue themselves to the solid substrate. A central molecule regulating the processes of this motile-sessile transition is the second messenger c-di-GMP, which stimulates the production of a variety of exopolysaccharide adhesins in different bacterial model organisms. In Caulobacter crescentus, c-di-GMP regulates the synthesis of the polar holdfast adhesin during the cell cycle, yet the molecular and cellular details of this control are currently unknown. Here we identify HfsK, a member of a versatile N-acetyltransferase family, as a novel c-di-GMP effector involved in holdfast biogenesis. Cells lacking HfsK form highly malleable holdfast structures with reduced adhesive strength that cannot support surface colonization. We present indirect evidence that HfsK modifies the polysaccharide component of holdfast to buttress its cohesive properties. HfsK is a soluble protein but associates with the cell membrane during most of the cell cycle. Coincident with peak c-di-GMP levels during the C. crescentus cell cycle, HfsK relocalizes to the cytosol in a c-di-GMP-dependent manner. Our results indicate that this c-di-GMP-mediated dynamic positioning controls HfsK activity, leading to its inactivation at high c-di-GMP levels. A short C-terminal extension is essential for the membrane association, c-di-GMP binding, and activity of HfsK. We propose a model in which c-di-GMP binding leads to the dispersal and inactivation of HfsK as part of holdfast biogenesis progression.IMPORTANCE Exopolysaccharide (EPS) adhesins are important determinants of bacterial surface colonization and biofilm formation. Biofilms are a major cause of chronic infections and are responsible for biofouling on water-exposed surfaces. To tackle these problems, it is essential to dissect the processes leading to surface colonization at the molecular and cellular levels. Here we describe a novel c-di-GMP effector, HfsK, that contributes to the cohesive properties and stability of the holdfast adhesin in C. crescentus We demonstrate for the first time that c-di-GMP, in addition to its role in the regulation of the rate of EPS production, also modulates the physicochemical properties of bacterial adhesins. By demonstrating how c-di-GMP coordinates the activity and subcellular localization of HfsK, we provide a novel understanding of the cellular processes involved in adhesin biogenesis control. Homologs of HfsK are found in representatives of different bacterial phyla, suggesting that they play important roles in various EPS synthesis systems.

On-line analysis of CAL72 cells on two different titanium surfaces in a perfusion micro-bioreactor.

OBJECTIVES: The aim of the present experiment was to test a prototype microsensoric measuring system (micro-bioreactor) for the investigation of the biocompatibility of different titanium surfaces in a cell culture model. METHODS: Osteosarcoma cells of the cell line CAL72 were seeded onto titanium plates (10mm x 10mm x 1mm) and inserted into the culture chamber of the micro-bioreactor. Titanium plates with two different surface topographies (machined and titanium plasma-sprayed [TPS]) were used for this pilot investigation. Plastic plates served as controls. The online-sensoric device of the micro-bioreactor allowed the continuous monitoring of the metabolism of the cells and the control of the culture conditions. Over a period of 17 h changes in O(2)-consumption in the medium were measured by micro-electrodes and registered by the software of the system. The metabolic activity of the cells was calculated from the difference between the bypass and the chamber values. The cell proliferation and vitality were analyzed before and after the perfusion time in the micro-bioreactor. The cell morphology was studied using scanning electron microscopy. RESULTS: The cells on the machined surfaces showed the highest oxygen consumption after 15 h, after that it decreased. The cells on the TPS plates showed a lower oxygen consumption, which remained stable after 17 h. The highest oxygen consumption was seen with the cells on the control plastic plates. Concerning cell proliferation analysis, it could be shown that more vital CAL72 cells seeded onto TPS and plastic could be detected after the passage through the micro-bioreactor. Hence, the number of vital cells on the machined surface was reduced after the passage. SIGNIFICANCE: Within the limits of this experiment, the presented micro-bioreactor system could offer a valuable method to examine the dynamic interactions of cells and materials under defined in vitro experimental conditions. While the presented system is already successfully used in the ecological/ecotoxicological field, its routine use for investigating dental materials on a cellular level has to be evaluated.