Structure and Protein-Protein Interactions of Ice Nucleation Proteins Drive Their Activity.

Microbially-produced ice nucleating proteins (INpro) are unique molecular structures with the highest known catalytic efficiency for ice formation. Airborne microorganisms utilize these proteins to enhance their survival by reducing their atmospheric residence times. INpro also have critical environmental effects including impacts on the atmospheric water cycle, through their role in cloud and precipitation formation, as well as frost damage on crops. INpro are ubiquitously present in the atmosphere where they are emitted from diverse terrestrial and marine environments. Even though bacterial genes encoding INpro have been discovered and sequenced decades ago, the details of how the INpro molecular structure and oligomerization foster their unique ice-nucleation activity remain elusive. Using machine-learning based software AlphaFold 2 and trRosetta, we obtained and analysed the first ab initio structural models of full length and truncated versions of bacterial INpro. The modeling revealed a novel beta-helix structure of the INpro central repeat domain responsible for ice nucleation activity. This domain consists of repeated stacks of two beta strands connected by two sharp turns. One beta-strand is decorated with a TxT amino acid sequence motif and the other strand has an SxL[T/I] motif. The core formed between the stacked beta helix-pairs is unusually polar and very distinct from previous INpro models. Using synchrotron radiation circular dichroism, we validated the ß-strand content of the central repeat domain in the model. Combining the structural model with functional studies of purified recombinant INpro, electron microscopy and modeling, we further demonstrate that the formation of dimers and higher-order oligomers is key to INpro activity. Using computational docking of the new INpro model based on rigid-body algorithms we could reproduce a previously proposed homodimer structure of the INpro CRD with an interface along a highly conserved tyrosine ladder and show that the dimer model agrees with our functional data. The parallel dimer structure creates a surface where the TxT motif of one monomer aligns with the SxL[T/I] motif of the other monomer widening the surface that interacts with water molecules and therefore enhancing the ice nucleation activity. This work presents a major advance in understanding the molecular foundation for bacterial ice-nucleation activity.

Terrestrial Origin for Abundant Riverine Nanoscale Ice-Nucleating Particles.

Ice-nucleating particles (INPs) associated with fresh waters are a neglected, but integral component of the water cycle. Abundant INPs were identified from surface waters of both the Maumee River and Lake Erie with ice nucleus spectra spanning a temperature range from -3 to -15 °C. The majority of river INPs were submicron in size and attributed to biogenic macromolecules, inferred from the denaturation of ice-nucleation activity by heat. In a watershed dominated by row-crop agriculture, higher concentrations of INPs were found in river samples compared to lake samples. Further, ice-nucleating temperatures differed between river and lake samples, which indicated different populations of INPs. Seasonal analysis of INPs that were active at warmer temperatures (≥-10 °C; INP-10) showed their concentration to correlate with river discharge, suggesting a watershed origin of these INPs. A terrestrial origin for INPs in the Maumee River was further supported by a correspondence between the ice-nucleation signatures of river INPs and INPs derived from the soil fungus Mortierella alpina. Aerosols derived from turbulence features in the river carry INP-10, although their potential influence on regional weather is unclear. INP-10 contained within aerosols generated from a weir spanning the river, ranged in concentration from 1 to 11 INP m-3, which represented a fold-change of 3.2 over average INP-10 concentrations sampled from aerosols at control locations.

Biomechanical patterns of text-message distraction.

The objective of this study was to identify biomechanical measures that can distinguish texting distraction in a laboratory-simulated driving environment. The goal would be to use this information to provide an intervention for risky driving behaviour. Sixteen subjects participated in this study. Three independent variables were tested: task (texting, visual targeting, weighted and non-weighted movements), task direction (front and side) and task distance (close and far). Dependent variables consisted of biomechanical moments, head displacement and the length of time to complete each task. Results revealed that the time to complete each task was higher for texting compared to other tasks. Peak moments during texting were only distinguishable from visual targeting. Peak head displacement and cumulative biomechanical exposure measures indicated that texting can be distinguished from other tasks. Therefore, it may be useful to take into account both temporal and biomechanical measures when considering warning systems to detect texting distraction.